CN102224165A - Transgenic plants with increased yield - Google Patents

Transgenic plants with increased yield Download PDF

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CN102224165A
CN102224165A CN2009801463883A CN200980146388A CN102224165A CN 102224165 A CN102224165 A CN 102224165A CN 2009801463883 A CN2009801463883 A CN 2009801463883A CN 200980146388 A CN200980146388 A CN 200980146388A CN 102224165 A CN102224165 A CN 102224165A
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amino acids
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B·D·麦克尔西
W·布鲁斯
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BASF Plant Science Co GmbH
BASF Plant Science GmbH
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Abstract

Polynucleotides are disclosed which are capable of enhancing yield of a plant transformed to contain such polynucleotides. Also provided are methods of using such polynucleotides and transgenic plants and agricultural products, including seeds, containing such polynucleotides as transgenes.

Description

Transgenic plant with output of increase
The application requires the U.S. Provisional Patent Application series number 61/115,947 of submission on November 19th, 2008; The U.S. Provisional Patent Application series number 61/107 that on October 23rd, 2008 submitted to, the U.S. Provisional Patent Application series number 61/099 that on September 23rd, 739 and 2008 submitted to, 224 benefit of priority, the complete content of every piece of patent documentation mode is by reference incorporated this paper into.
Invention field
Present invention relates in general to the transgenic plant of the isolating polynucleotide of overexpression coded polypeptide in specified plant tissue and organoid, thereby improve the output of described plant.
Background of invention
Population increases and climate change may make global food, feed and fuel crunch become outstanding focal issue in recent years.Agricultural has consumed 70% of water that the people uses, and the mass part of rainfall this moment in the world continues to descend.In addition, along with land use becomes city and suburb from the farmland, still less Gong Qing arable land can be used for cultivating agricultural crops.Agricultural biotechnologies have attempted modifying to satisfy human growing demand by the plant genetic that may improve crop yield, for example reply tolerance or pass through to increase biomass by giving better abiotic stress.
Crop yield is defined as the bushel number of every acre of relevant agricultural-food of being gathered in the crops (as grain, feed or seed) in this article.Crop yield is subjected to abiotic stress as arid, heat, salt with coldly coerce and be subjected to plant size (biomass) influence.Conventional plant breeding strategy is successfully not give the abiotic stress tolerance of increase relatively slowly and generally.Improve grain yield by the conventional breeding method and in corn, almost reached a plateau.Harvest corn index (that is, during results output biomass to the ratio of total accumulation biomass) basically during going through nearest hundreds of years grain yield selection breeding and has not changed.Therefore, the recent output that has taken place in corn is improved the result of the total biomass generation that is the increase of per unit land area.By increasing the total biomass that planting density realizes this increase, wherein said increase planting density has caused the adaptability phenotypic alternation, as reducing the fringe size that the leaf angle (leaf angle) of covering the bottom leaf reduces and can improve harvest index.
If when the soil water runs out or water be unavailable at arid time durations, then crop yield is limited.If the transpiration of leaf surpasses the water supply from root, then plant hydropenia occurs.Amount and the plant of holding water in the supply of available water and the soil are relevant with the ability that its root system system contacts this water.Water leaves the transpiration of leaf and passes through photosynthetic fixing relevant through air passing hole with carbonic acid gas.These two processes are positively related, thereby closely related with the water loss due to transpiration through photosynthetic high carbon dioxide influx.Along with water is rising from leaf, leaf water potential reduction and pore are inclined in the hydraulic pressure process of the photosynthetic amount of restriction and close.Because the fixing of carbonic acid gas in the photosynthesis depended in crop yield, so water intake and transpiration are the contribution factors of crop yield.Can use water still less to fix the carbonic acid gas of same amount or can have the more photosynthesis of enforcement the plant that the low flow of water runs well and thereby in many agrosystems, produce the more potential of multi-biomass and economic yield.
Agricultural biotechnologies scholar has used most its of the assay method in model plant system, the research of crop plants greenhouse and the field test to develop the transgenic plant that increase or show because of the biomass increase output increase because of abiotic stress tolerance hardy.For example, water application efficiency (WUE) often is a parameter relevant with drought tolerance.The research of plant being replied drying, osmotic shock and extreme temperature also is used for determining tolerance or the resistance of this plant at abiotic stress.
The increase of biomass can be owing to the water consumption of growth efficiency that improves relatively or minimizing under low water operability.Selecting in the proterties for the improvement crop, under the unconverted situation of growth, the reduction meeting that water uses has peculiar advantage in water input Irrigation farming system with high costs.Do not have under the situation of corresponding input in the water use, growth increases the suitability that will have at whole agrosystems.In the many agrosystems that do not limit the water supply, growth increases and also increases output, even if this increase is with the cost that increases to of water use.
Agricultural biotechnologies scholar also uses the value of other parameters that show the crop yield of transgenosis potential impact.For fodder crop such as clover, silage corn and hay, phytomass is relevant with ultimate production.Yet for cereal crop, used other parameters to come estimated output, as the plant size, it is long-pending by total plant dry weight, over-ground part dry weight, over-ground part fresh weight, leaf area, caulome, plant height, rosette diameter, leaf length, root length, root quality, tiller number and number of sheets tolerance.The plant size of growing commitment general with grow in after a while plant big or small relevant.Have plant absorbing more rays that the big plant of bigger leaf area generally can be smaller and carbonic acid gas and thereby may be during the identical period deserved bigger weight.Existence is at the strong hereditary component of plant size and growth velocity, and thereby with regard to a series of different genetic phenotypes, relevant with size under the another kind of envrionment conditions probably in the plant size under a kind of envrionment conditions.By this way, use standard environment to simulate field crops in different positions and difference that is met with on the time and dynamic environment.
Harvest index is metastable under many envrionment conditionss, and thereby plant size and grain yield between sane relevant be possible.Plant size and grain yield are internal associations, because most cereal biomass depends on the photosynthetic productivity of the existing or deposit of leaf and stem.As for the abiotic stress tolerance, measuring the plant size that is in early development under the normalization condition in growth room or greenhouse is to measure the standard practices that is had the potential production advantage of giving by transgenosis.
Prescribe a time limit when the water operability has, the vegetable cell protein called membrane transporters are often influenced.Under extreme case, water shifting out from this cytolemma destroyed normal bilayer structure and cause this film unusual porous that becomes when being dried.Under the condition of milder, can cause protein called membrane transporters displacement and configuration thereof to change to coercing of double-layer of lipoid, cause lower molecule transport efficacy.Olighydria also can improve cytosol concentration, and this transfers to influence the protein configuration of (comprising translocator).
Health, the g and D of crop plants under the varying environment condition depended in crop yield.The correct targeted delivery of mineral nutrient and organic compound and in time send to plant-growth and grow essential.Stress conditions such as the arid normal delivery system in can the havoc plant.Under this type of stress conditions, make the stable gene of molecule transhipment help to keep stable state in the plant.
The molecule transhipment of being regulated needs energy to be used for many processes of plant.The ion gradient of cross-cell membrane and proton gradient are a kind of forms of energy reserve in the vegetable cell.These gradients are used for driving other molecule transmembrane transports.An example is the plastosome electron transport chain, and it uses the reduction energy of NADH to come cross-line plastochondria inner membrance to move proton, thereby produces the gradient of pH and electric charge.Another example is the electron transport chain in the chloroplast(id), and it becomes photosynthesis may be so that produce proton gradient of striding thylakoid membrane and the reducing power that produces the NADPH form with photon energy.In both cases, become the chemical energy of ATP form by film mating type ATP enzymatic conversion from the energy in the proton gradient (being called proton motive force) of cross-line plastochondria film or thylakoid membrane.Main active transport is by the energy that directly uses in this transport process from ATP that acts on of ATP enzyme, and wherein said ATP enzyme cuts the terminal phosphate of ATP, thereby forms ADP.
The ATP enzyme is that a class catalysis ATP resolves into ADP and free phosphoric acid salt ion or catalysis reversed reaction to produce the enzyme of ATP.The dephosphorylation reaction releases energy, and this energy is used for striding film and moves solute.Stride toxin, refuse and solute that essential many metabolites of film ATP enzyme input cellular metabolism and output may hinder cell processes.Except the exchange material, the film ATP enzyme of striding of other types comprises cotransporter and pump.
The ATP enzyme can be different aspect function, structure and the ionic type of transporting at their.F-ATP enzyme in mitochondrial membrane, chloroplast membranes and the bacterial membrane is the main producer of ATP, and it uses the proton gradient by photosynthesis produced in oxidative phosphorylation in the plastosome or the chloroplast(id).The A-ATP enzyme is present in the ancient bacterium (Archaea) and as the F-ATP enzyme and plays a role.The V-ATP enzyme mainly is present in the eukaryote vacuole, and catalysis ATP hydrolysis is transported solute and reduced pH in the organoid.The V-ATP enzyme plays a role as proton pump exclusively.Use the proton motive force that produces by V-ATP enzyme in eukaryotic organoid and the film motivating force subsequently as many secondary transport processes.The P-ATP enzyme is present in bacterium, fungi and is present in Eukaryotic after birth and the organoid, and the effect of the multiple different ions of performance transmembrane transport.The E-ATP enzyme is a series of NTP of the hydrolysis ectocellular enzymes of (comprising the outer ATP of born of the same parents).
Opposite with elementary active transport, secondary active transport uses the energy from concentration gradient, and wherein said concentration gradient is set up in advance by above-mentioned process.There is 2 types secondary active transport process: exchange transhipment (antiport) and co-transport (symport).Amino acid and sugar transport are undertaken by secondary active transport mechanism.
ABC (ATP-binding cassette) translocator is a transmembrane protein, and the substrate that it utilizes ATP hydrolysis energy to stride epicyte and stride intracellular membrane transhipment wide range of types comprises meta-bolites, lipid and sterol and medicine.In bacterium, the essential compound of the main pumping of abc transport albumen as sugar, VITAMIN and metal ion to cell.In eukaryote, abc transport albumen mainly is transported to molecule the outside of after birth or to film mating type organoid such as endoplasmic reticulum and plastosome.
Electrotransfer reaction is basic for the main energy metabolism in plant mitochondria (respiration) and the chloroplast(id) (photosynthesis).In these two kinds of organoids, electronics has produced the proton motive force of striding film from another molecule that a molecule on cytolemma one side is passed on this film offside.Though efficient, the electronics that the electron transfer process in plant mitochondria and chloroplast(id) spills little percentage ratio comes partly oxygen reduction, thereby form active oxygen such as super-oxide.Cellular energy is not only wasted in the formation of super-oxide, also may cause the oxidative stress that promotes that cell function descends because of damage membrane lipid, protein and DNA.In addition, existence from light harvesting complex the activated chlorophyll molecule to the transmission ofenergy of molecular state triplet oxygen (molecular triplet oxygen) with form singlet oxygen (singlet oxygen) may, wherein said singlet oxygen is the another kind of precursor of active oxygen species.The trend of photosystem and light harvesting complex excited oxygen improves or reduces the blocking effect that the substrate level exceeds critical threshold in because of the eubolism approach and increase coercing time durations.
Respiration in the plant mitochondria can be transferred to adenosine triphosphate (ATP) by the biological chemistry that a series of katabolism redox reactions will be derived from nutrient substance.Generally speaking, sugar and amino acid and lipid acid use the energy that discharges to synthesize ATP as transmitting the substrate of electronics to oxygen.General reaction for sugar can be reduced to C 6H 12O 6+ 6O 2→ 6CO 2+ 6H 2O accompanies by Δ Hc-2880kJ.In plant mitochondria, tricarboxylic acid cycle reaction discharges and is used to reduce NAD and becomes the electronics of NADH.Redox energy from NADH is passed to oxygen by electron transport chain.Electronics has discharged along this transmission of inner membrane protein matter complex body and has produced the energy of striding membranous sub-gradient.The gained proton motive force of cross-line plastochondria film is used for synthesizing ATP.The energy that is stored among the ATP is used for a plurality of cell processes that need energy, comprises the cross-cell membrane transhipment of biosynthesizing and molecule.
Photosynthesis is that the bacterium of plant and some type is used from the energy of sunlight so as to from carbonic acid gas (CO 2) and water in produce the complex process of glucose and oxygen.Overall chemical reaction can be expressed as 6CO simply 2+ 6H 2O (+luminous energy) → C 6H 12O 6+ 6O 2The many reactions that during photosynthesis, take place be divided into usually 2 the stage-inner and transmit " photoresponse " of electronics and proton and relate at photosynthetic membrane from CO across photosynthetic membrane 2" dark reaction " of biosynthesizing carbohydrate.Use is arranged in the subunit photosystem more than two kinds (I and II) of the thylakoid membrane of chloroplast(id), and higher plant catches luminous energy.This electron transport produces the proton gradient of striding thylakoid membrane, and wherein the described proton gradient of Chan Shenging is used for the synthetic of ATP.Photoresponse in the photosynthesis produces ATP and NADPH, and the two is used for producing the biochemical reaction of sugar, amino acid and other cellular components subsequently.
Photosystem I (PS-I) is many subunits complex body, and it uses luminous energy to drive the electronics of being supplied with from photosystem II (PSII) and strides the thylakoid membrane transhipment and become NADPH with reduction NADP.The electron transport of PS-I catalysis from plastocyanin (it is positioned at the side, chamber of thylakoid) to the optical drive of ferredoxin (it is on matter side between this film).PS-I complex body heart place therein has the PsaA/PsaB heterodimer, and this heterodimer contains the chlorophyll dimer of main electron donor-be called P700-and electron acceptor(EA) A0, A1 and FX/A/B.Numerous less protein subunits constitute the rest part of this complex body.Some subunits in these subunits serve as the binding site of soluble electron carrier plastocyanin and ferredoxin, although also do not know better some proteinic functions in all the other protein.Huge antenna system with about 90 chlorophyll and 22 carotenoid is caught light and is shifted excitation energy to this center.P700 uses the electronics of sending from PS-II to restore by plastocyanin.PsaF promotes plastocyanin or cytochrome c (being responsible for the movable electron carrier of reduction-oxidation type donor P700) bonded plastocyanin docking protein among the PS-I.U.S. Patent Application Publication 2008/0148432 has disclosed the purposes of agronomy proterties in the PS-I PsaF gene enhancing transgenic plant.
PS-II (also be contain photosynthetic membrane be the oligomeric protein-pigment complex body of intrinsic and external polypeptide) uses luminous energy to come oxidizing water.PS-II has the P680 reactive center that contains chlorophyll a.Core inside at this complex body, chlorophyll and β-Hu Luobusu pigment mainly combine with protein C P43 (PsbC) and CP47 (PsbB), described protein is sent to reactive center protein D 1 (Qb with excitation energy, PsbA) and D2 (Qa, PsbD), wherein said reactive center protein binding participates in whole redox active cofactors of energy conversion process.PS-II is put oxygen complex body (oxygen-evolving complex; OEC) oxidizing water to be providing the proton that is used by PS-I, and is made up of OEE1 (PsbO), OEE2 (PsbP) and OEE3 (PsbQ).Residue subunit among the PS-II is low-molecular-weight (less than 10kDa), and participates in PS-II assembling, stabilization, removes singletization (demonization) and photo-protection.PsbW is the part of this lower molecular weight transmembrane protein complex body, and it is this subunit of putting the oxygen complex body therein.As if PsbW has several effects, comprises that the PS-II after instructing the PS-II biology to generate and assemble, make dimer PS-II stabilization and promoting the light restraining effect repairs.U.S. Patent Application Publication 2007/0067865 has disclosed the conversion plant with nucleic acid molecule, and it can be the structural nucleic acid of PsbW gene that wherein said nucleic acid molecule comprises.
Electronics from photosystem is transferred to molecular oxygen once in a while, thereby forms super-oxide, a kind of precursor of more activated oxygen intermediate.One of key point of this transfer is at the ferredoxin place.Ferredoxin is omnipresence [2Fe-2S] protein of many electron transport routes in involved in plant, animal and the microorganism.Ferredoxin (PetF) is the electron carrier protein in the PS-I electron transport chain.In this chain, ferredoxin transhipment from the electronics of PS-I to ferredoxin-NADP oxydo-reductase, its catalysis from the electron transport of Fd to NADP+ to produce NADPH.In addition, the reductibility equivalent from ferredoxin is used for nitrogen and sulfur assimilation and amino acid and fatty acid metabolism.Ferredoxin also provides and has been used for the reductibility equivalent that Trx activates chloroplast enzyme.Think that high-caliber ferredoxin is crucial for the survival of the plant in the suboptimum environment.In higher plant, ferredoxin is by minigene family coding, the expression that described minigene family has tissue specificity and regulated by environment.The gene of coding ferredoxin is owing to sideropenia, oxidative stress and serious environment-stress (comprising arid, cold, salinity and ultraviolet ray) are reduced.The amount of ferredoxin mRNA is subjected to the redox modulating on the post-transcriptional level, and use transgenic method increase plant iron oxygen also protein gene expression and improve the strategy of stress tolerance in the crop thereby success.Plastosome also contains the ferredoxin that participates in electrotransfer reaction.
Flavodoxin has redox-potential and the function similar with the ferredoxin in the algae to cyanobacteria, but its gene is not present in any Plant Genome.Flavodoxin participates in the development of stress tolerance in cyanobacteria and the algae.U.S. Patent number 6,781,034 expression of flavodoxin gene in tobacco that disclose from anabena (Anabaena) produce the transgenic plant with the arid of increase, high intensity of illumination, heat, cold, UV radiation and herbicides paraquat tolerance.
Chlorophyll is the main ingredient around the light harvesting complex of photosystem I and II (light harvesting complex).It structurally is similar to other porphyrin pigments such as protoheme and produces by identical with it pathways metabolism.Be magnesium ion at the center of this ring and connect different side chains, generally include long phytol chain.Cobalami is to share the complicated small molecules that is produced in the approach of commitment by microorganism with the chlorophyll biosynthetic pathway exclusively.Cobalami approach and chlorophyll approach all are derived from common precursor, uroporphyrinogen III.The complicacy of cobalami (vitamin B12) itself and singularity and generation thereof require about 30 kinds of enzymes, and described enzyme picks out special but closely-related substrate in the chemically complicated approach.A kind of enzyme like this (uroporphyrin-III C-methyltransgerase) two successive C-2 of catalysis and C-7 methylation reaction, wherein said methylation reaction participation uroporphyrinogen-III changes into preceding corrin-2 by the intermediate forms of preceding corrin-1.This reaction instructs uroporphyrinogen-III to enter cobalami (vitamin B12) or helicorubrin (siroheme) biosynthesizing.U.S. Patent Application Publication 2005/0108791 has disclosed cytoalgae species (Synechocystis sp.) uroporphyrin III C-methyltransgerases (CobA) generation with chlorophyll targeted peptide and has had the purposes of the transgenic plant of improving phenotype.
Characterized some genes that relate to stress response in the plant, water use and/or biomass, but so far, limited ground successfully exploitation has the genetically modified crops plant of improving output, and all not commercializations of this type of plant.Therefore, need to identify extra gene with the ability that increases crop plants output.
Summary of the invention
The inventor has been found that when the subcellular location that some polynucleotide is expressed on suitable level and the protein target of gained is suitable, causes the improvement of plant biomass with described gene-transformed plant.When target as described herein, polynucleotide described in the table 1 and polypeptide can improve the output of transgenic plant.
Table 1
Figure BDA0000062684420000081
Figure BDA0000062684420000091
In one embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; With the isolating polynucleotide of coding full-length polypeptide, described full-length polypeptide has and is selected from the NO:2 by SEQ ID; SEQ ID NO:4; SEQ ID NO:6; Sequence in the group of forming with SEQ ID NO:8; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; With the isolating polynucleotide of coding chloroplast transit peptides and the isolating polynucleotide of coding full-length polypeptide, described full-length polypeptide has the sequence that is selected from the group of being made up of SEQ ID NO:10 and SEQ ID NO:12; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of coding line plastochondria transit peptides; Have the isolating polynucleotide of 11 possible isoprene pyrophosphate synthetase polypeptide of the total length of sequence described in SEQ ID NO:14 with coding; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides and use the expression cassette transgenic plant transformed, described expression cassette comprises the coding that is in effective connection can strengthen the separation polynucleotide of promotor of genetic expression in the leaf and the isolating polynucleotide of coding line plastochondria transit peptides; With the isolating polynucleotide of coding full-length polypeptide, described full-length polypeptide is the transcriptional of inferring with fatty acid metabolism of the gntR type HTH DNA binding domains that comprises SEQ ID NO:16 the 34th to the 53rd amino acids; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; Has G3E with coding, the isolating polynucleotide of the full-length polypeptide in P ring structure territory, described G3E, P ring structure territory has comprised Walker A motif with sequence described in SEQ ID NO:99 and the GTP specificity motif with sequence described in SEQ ID NO:100; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides and use the expression cassette transgenic plant transformed, described expression cassette comprises the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf, with the isolating polynucleotide of coding full-length polypeptide, described full-length polypeptide is the membranin of inferring with sequence described in SEQ ID NO:22; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; Isolating polynucleotide with coding line plastochondria transit peptides; Comprise the isolating polynucleotide of the total length peroxysome-coenzyme A synthetic enzyme polypeptide of AMP binding domains with coding, described AMP binding domains is selected from the 194th to the 205th amino acids by SEQ ID NO:24, SEQ ID NO:26 the 202nd to the 213rd amino acids, SEQ ID NO:28 the 214th to the 225th amino acids, SEQ ID NO:30 the 195th to the 206th amino acids, SEQ ID NO:32 the 175th to the 186th amino acids, SEQ ID NO:34 the 171st to the 182nd amino acids, SEQ ID NO:36 the 189th to the 200th amino acids, the group that SEQ ID NO:38 the 201st to the 212nd amino acids is formed; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; Isolating polynucleotide with coding line plastochondria transit peptides; Have the isolating polynucleotide of the total length histone H 4 polypeptide of G-A-K-R-H (SEQ ID NO:101) characteristic sequence structural domain with coding, described G-A-K-R-H characteristic sequence structural domain is selected from the 3rd to the 92nd amino acids by SEQ ID NO:40; SEQ ID NO:56 the 3rd to the 92nd amino acids; The group that SEQ ID NO:42 the 3rd to the 92nd amino acids and SEQ ID NO:44 the 3rd to the 92nd amino acids are formed; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides use expression cassette transgenic plant transformed, described expression cassette comprise the coding that is in effective connection can strengthen genetic expression in the leaf promotor or the coding constitutive promoter the separation polynucleotide; The isolating polynucleotide of the isolating polynucleotide of coding chloroplast transit peptides and coding total length SYM1 type conformity membrane polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of the isolating polynucleotide of coding line plastochondria transit peptides and coding total length vacuolar proton pump subunit H polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding line plastochondria transit peptides; Comprise the isolating polynucleotide of the total length F-ATP enzyme subunit α polypeptide of atp synthase structural domain with coding, described atp synthase structural domain is selected from the 356th to the 365th amino acids by SEQ ID NO:62; The group that SEQ ID NO:64 the 254th to the 263rd amino acids is formed; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding line plastochondria transit peptides; Comprise the isolating polynucleotide of the total length F-ATP enzyme subunit beta polypeptides of atp synthase structural domain with coding, described atp synthase structural domain is selected from the group of being made up of SEQ ID NO:66 the 353rd to the 362nd amino acids; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding line plastochondria transit peptides; Have the isolating polynucleotide of the total length abc transport protein polypeptide of sequence described in SEQ ID NO:68 with coding; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding plastid transit peptides; Have the isolating polynucleotide of the total length photosystem I reactive center subunit psaK polypeptide of psaGK label with coding, described psaGK label comprises the 56th to the 73rd amino acids of SEQ ID NO:70; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding line plastochondria transit peptides and coding comprise the isolating polynucleotide of the total length ferredoxin polypeptide of Fer2 characteristic sequence, and described Fer2 characteristic sequence is selected from the 11st to the 87th amino acids by SEQ ID NO:72; The group that SEQ ID NO:74 the 12nd to the 88th amino acids and SEQ ID NO:76 the 63rd to the 139th amino acids are formed; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding plastid transit peptides; Have the isolating polynucleotide of the total length flavodoxin polypeptide of flavodoxin _ 1 characteristic sequence with coding, described flavodoxin _ 1 characteristic sequence comprises the 6th to the 160th amino acids of SEQ ID NO:78; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding plastid transit peptides; Comprise the isolating polynucleotide of the total length photosystem I reactive center subunit III psaF polypeptide of PSI_PsaF characteristic sequence with coding, described PSI_PsaF characteristic sequence is selected from the 3rd to the 158th amino acids by SEQ ID NO:80; SEQ ID NO:82 the 43rd to the 217th amino acids; SEQ ID NO:84 the 46th to the 220th amino acids; SEQ ID NO:86 the 50th to the 224th amino acids; Group with SEQ ID NO:88 the 50th to the 224th amino acids composition; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding line plastochondria transit peptides; Have the isolating polynucleotide of total length cytochrome c 553 (PetJ) polypeptide of PSI_PsaF characteristic sequence with coding, described PSI_PsaF characteristic sequence comprises the 38th to the 116th amino acids of SEQ ID NO:90; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding line plastochondria transit peptides; Have the isolating polynucleotide of total length photosystem II reactive center W (PsbW) polypeptide of cytochrome C characteristic sequence with coding, described cytochrome C characteristic sequence comprises the 5th to the 120th amino acids of SEQ ID NO:92; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding plastid transit peptides; Have the isolating polynucleotide of total length uroporphyrin-III c-methyltransgerase (CobA) polypeptide of sequence described in SEQ ID NO:93 with coding; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
In another embodiment, the invention provides and use the expression cassette transgenic plant transformed, described expression cassette comprises the isolating polynucleotide of the isolating polynucleotide of the coding promotor that is in effective connection and the preceding corrin of total length-6b methylase that coding has methyltransgerase _ 12 characteristic sequences, and described methyltransgerase _ 12 characteristic sequences comprise the 45th to the 138th amino acids of SEQ ID NO:96; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.The expression cassette of this embodiment can randomly comprise the isolating polynucleotide of coding line plastochondria transit peptides.
In another embodiment, the invention provides and use the expression cassette transgenic plant transformed, described expression cassette comprises the isolating polynucleotide of the isolating polynucleotide of the coding promotor that is in effective connection and the preceding corrin of decarboxylation-6y methylase that coding has TP_ methylase characteristic sequence, and described TP_ methylase characteristic sequence comprises the 1st to the 195th amino acids of SEQ ID NO:98; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.The expression cassette of this embodiment can randomly comprise the isolating polynucleotide of coding line plastochondria transit peptides.
In another embodiment, the invention provides the seed that produces by transgenic plant of the present invention, wherein seed isozygotys for the transgenosis that comprises expression vector mentioned above.The plant that is derived from seed of the present invention is compared environmental stress-tolerance and/or the plant-growth of increase and/or the output of increase of demonstration increase with the wild-type kind of this plant under normal or stress conditions.
In yet another aspect, the present invention relates to by or from transgenic plant of the present invention, their plant part or the product that produces of its seed, as food, feed, food fill-in, feed supplement, fiber, makeup or medicine.
The present invention also is provided in the table 1 some isolating polynucleotide of being identified and some isolated polypeptide of being identified in table 1.The present invention also is embodied in the recombinant vectors that comprises isolating polynucleotide of the present invention.
In another embodiment, the present invention relates to produce the method for aforementioned transgenic plant, wherein this method comprises with the expression vector transformed plant cells that comprises isolating polynucleotide of the present invention, and from these vegetable cell generation transgenic plant, wherein said transgenic plant are expressed the polypeptide by described polynucleotide encoding.Compare with the wild-type kind of this plant, the expression of described polypeptide in this plant causes normally and/or the environmental stress-tolerance that increases under the stress conditions and/or the growth and/or the output of increase.
In another embodiment, the invention provides the environmental stress-tolerance of increase plant and/or the method for growth and/or output.Described method comprises step: with the expression cassette transformed plant cells that comprises isolating polynucleotide of the present invention, and from these vegetable cell generation transgenic plant, wherein said transgenic plant comprise this polynucleotide.
The accompanying drawing summary
Fig. 1 shows the comparison result of the proteinic aminoacid sequence that contains the Nucleotide binding domains, and described protein is called B2173 (SEQ ID NO:18), GM50181105 (SEQ ID NO:20).Use the Align X of Vector NTI to produce this comparison result.
Fig. 2 shows the comparison result of the aminoacid sequence of the peroxysome-coenzyme A synthetic enzyme that is called YBR222C (SEQ ID NO:24), BN51408632 (SEQ ID NO:26), BN51423788 (SEQ ID NO:28), BN51486050 (SEQ ID NO:30), GM50942269 (SEQ ID NO:32), GM59534234 (SEQ ID NO:34), GM59654631 (SEQ ID NO:36), GM59778298 (SEQ ID NO:38).Use the Align X of Vector NTI to produce this comparison result.
Fig. 3 shows the comparison result of the aminoacid sequence of the histone H 4 that is called YNL030W (SEQ ID NO:40), GM53663330 (SEQ ID NO:56), LU62237699 (SEQ ID NO:42), OS36075085 (SEQ ID NO:44).Use the Align X of Vector NTI to produce this comparison result.
Fig. 4 shows the comparison result of the proteic aminoacid sequence of SYM1 type conformity membrane that is called YLR251W (SEQ ID NO:62), BN42108421 (SEQ ID NO:64), GMsf23a01 (SEQ ID NO:50), HV62697288 (SEQ ID NO:52), LU61649286 (SEQ ID NO:54), OS40298410 (SEQ ID NO:56).Use the Align X of Vector NTI to produce this comparison result.
Fig. 5 shows the comparison result of the V-ATP enzyme subunit H amino acid sequence of polypeptide that is called YPR036W (SEQ ID NO:58), BN51362135 (SEQ ID NO:60).Use the Align X of Vector NTI to produce this comparison result.
Fig. 6 shows the comparison result of the aminoacid sequence of the F-ATP enzyme subunit α that is called SLL1326 (SEQ ID NO:62), LU61815688 (SEQ ID NO:64).Use the Align X of Vector NTI to produce this comparison result.
Fig. 7 shows the comparison result of the aminoacid sequence of the ferredoxin that is called sll1382 (SEQ ID NO:72), BN42448747 (SEQ ID NO:74), GM49779037 (SEQ ID NO:76).Use the Align X of Vector NTI to produce this comparison result.
Fig. 8 shows the comparison result of the proteic aminoacid sequence of photosystem I reactive center subunit III that is called sll0819 (SEQ ID NO:80), BN51362302 (SEQ ID NO:82), BNDLM1779_30 (SEQ ID NO:84), GMsk95f02 (SEQ ID NO:86) and GMso56a01 (SEQ ID NO:88).Use the Align X of Vector NTI to produce this comparison result.
Description of Preferred Embodiments
In the full text scope of the application's book, with reference to multiple publication.All the disclosure thereby the mode by reference of complete those reference of quoting incorporated the application's book in these publications and these publications, is intended to describe more fully prior art state related to the present invention.The purpose of used term only is to describe specific embodiments herein, and to be not intended to be restrictive.As used herein, " one " or " one " can mean one or more, and this depends on the context that uses this article.Therefore, for example, can mean the denotion of " cell " and can use at least one cell.
In one embodiment, the invention provides transgenic plant, the isolating polynucleotide that it is identified in overexpression table 1 in subcellular compartment shown in this article and tissue.When transgenic plant of the present invention are compared with the wild-type kind of this plant, show the output of improving.As used herein, term " output of improvement " means any improvement on the output of any measured plant product (as grain, fruit or fiber).According to the present invention, the variation of different phenotypic characters can improve output.For example and not limit, parameter such as development of floral organs, root are made a start, root biomass, seed number, seed weight, harvest index, abiotic environment stress tolerance, leaf one-tenth, phototropism, apical dominance and fruit development are to improve the suitable of output to measure.Any increase of output is the output according to improvement of the present invention.For example, the improvement of output can comprise that any measured parameter improves 0.1%, 0.5%, 1%, 3%, 5%, 10%, 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more.For example, when comparing with the bushel/acre yield of untreated soybean of cultivating down from the same terms or cereal, from the bushel/acre yield increase of the crop institute deutero-soybean that comprises following plant or cereal is output according to improvement of the present invention, and wherein said plant is genetically modified for the Nucleotide and the polypeptide of table 1.
As defined herein, " transgenic plant " are to have used the recombinant DNA technology change containing the plant of isolating nucleic acid, wherein said isolating nucleic acid otherwise can not be present in this plant.As used herein, term " plant " comprises complete plant, vegetable cell and plant part.Plant part includes but not limited to stem, root, ovule, stamen, leaf, embryo, meristem zone, callus, gametophyte, sporophyte, pollen, sporule etc.Transgenic plant of the present invention can be male sterile or male fertile, and can also comprise such transgenosis, and it is different from and comprises those transgenosiss of described isolating polynucleotide herein.
As used herein, term " kind " refers to such one group of plant in the species, and described plant is total to make them be different from the constant characteristic that common form in these species and other may kinds.When having at least a obvious proterties, a kind is a feature with some variations between all individualities in this kind also, and this mainly separates based on the Mendelian of proterties in the middle of the filial generation of follow-up generation.Think that a kind is " isozygotying " for a specific trait, if this kind is isozygotied in heredity in so much to this specific trait, so that when this isozygotys kind oneself pollination, in the middle of filial generation, do not observe the independent separate of the obvious amount of this proterties.In the present invention, this proterties produces because of the one or more isolating polynucleotide that the expression of transgenosis ground imports the plant product.Also as used herein, term " wild-type kind " is pointed out in comparing purpose one group of analyzed plant of plant in contrast, wherein said wild-type product kind of plant is identical with these transgenic plant (with isolating polynucleotide plant transformed of the present invention), except this wild-type product kind of plant isolating polynucleotide of the present invention of no use transform.Term " wild-type " refers to not do with isolating polynucleotide of the present invention vegetable cell, seed, plant component, plant tissue, plant organ or the complete plant of genetic modification as used in this article.
Term " control plant " refers to be used for relatively to be intended to identify enhanced phenotype in the plant of this transgenosis or genetic modification with the plant of transgenosis or genetic modification or vegetable cell, explant, seed, plant component, plant tissue, plant organ or the complete plant of the proterties wanted as used in this article.In some cases, " control plant " can be the transgenic plant strain, and it comprises empty carrier or marker gene, is not present in the transgenosis estimated or the purpose recombination of polynucleotide in the genetically modified plant but do not contain.Control plant can be the plant of identical strain or kind with the plant of tested genetically modified or genetic modification, or it can be other strain or kind, as known plant with particular phenotype, feature or known type.Suitable control plant comprises change or not genetically modified plant in the heredity that is used for producing the parental line of transgenic plant herein.
As defined herein, term " nucleic acid " and " polynucleotide " are tradable mutually and refer to the RNA or the DNA of linearity or ramose, strand or doubly-linked, or its crossbred.This term also comprises the RNA/DNA crossbred." isolating " nucleic acid molecule is a kind of nucleic acid molecule that separates basically with other nucleic acid molecule (sequences of other polypeptide of promptly encoding) that are present in this nucleic acid natural origin.For example, think that the nucleic acid of cloning is isolating.If nucleic acid has changed or has been placed in because of human intervention is not in the locus or position at this natural position of nucleic acid, if or it by the conversion method transfered cell, think also that then this nucleic acid is isolating.In addition, isolated nucleic acid molecule (as the cDNA molecule) can not contain and natural some related other cellular material of this nucleic acid molecule, maybe when producing by recombinant technology, does not contain substratum, or when chemosynthesis, does not contain precursor or other chemical.Isolated nucleic acid molecule can randomly comprise the non-translated sequence of 3 ' and the 5 ' end that is positioned at certain gene coding region, yet, may preferably remove in the naturally occurring replicon of this nucleic acid molecule natural distributed in the sequence of described coding region flank.
As used herein, term " environment-stress " refers to the suboptimum condition relevant with salinity, arid, nitrogen, temperature, metal, chemical, pathogenic agent or oxidative stress or its arbitrary combination.As used herein, term " arid " refers to wherein can be used for supporting the envrionment conditions of the water yield of plant-growth or growth less than the best.As used herein, term " fresh weight " refers to every kind of things in the plant, comprises water.As used herein, the total material outside term " dry weight " refers to dewater in the plant, and comprise for example carbohydrate, protein, oils and mineral nutrient.
Can transform arbitrarily plant species to produce transgenic plant of the present invention.Transgenic plant of the present invention can be dicotyledons or monocotyledons.For example and not limit, transgenic plant of the present invention can be derived from following any dicotyledons section: pulse family (Leguminosae) comprises plant such as pea, clover and soybean; Umbelliferae (Umbelliferae) comprises plant such as Radix Dauci Sativae and celery (celery); Solanaceae comprises plant such as tomato, potato, eggplant (aubergine), tobacco and capsicum; Cruciferae (Cruciferae), especially Btassica (Brassica) comprises plant such as oilseed rape, beet, Caulis et Folium Brassicae capitatae, Cauliflower (cauliflower) and blue and white cabbage (broccoli) and Arabidopis thaliana (A.thaliana); Composite family (Compositae) comprises plant such as lettuce (lettuce); Malvaceae (Malvaceae) comprises plant such as cotton; And Papilionaceae (Fabaceae), comprise plant such as Semen arachidis hypogaeae (peanut) etc.Transgenic plant of the present invention can be derived from monocotyledons, for example, and wheat, barley, Chinese sorghum, millet, rye, triticale (triticale), corn, rice, oat and sugarcane.Transgenic plant of the present invention also are presented as tree, for example apple tree, pear tree, pawpaw are set (quince), Japanese plum, cherry tree, peach, nectarine tree (nectarine), apricot, papaya (papaya), mango and other woody species, comprise softwood tree and deciduous trees, as willow (poplar), pine (pine), Chinese larch (sequoia), cdear (cedar), Oak Tree (oak) etc.Particularly preferably be Arabidopis thaliana, tobacco (Nicotiana tabacum), rice, oilseed rape, canola oil dish, soybean, cereal (corn) (corn (maize)), cotton and wheat.
A. the protein that does not characterize of non-target
In one embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; With the isolating polynucleotide of coding full-length polypeptide, described full-length polypeptide has and is selected from the NO:2 by SEQ ID, SEQ ID NO:4; SEQ ID NO:6; Sequence in the group of forming with SEQ ID NO:8; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
B. the agnoprotein matter of plastid target
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; With the isolating polynucleotide of coding chloroplast transit peptides and the isolating polynucleotide of coding full-length polypeptide, described full-length polypeptide has the sequence that is selected from the group of being made up of SEQ ID NO:10 and SEQ ID NO:12; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
C. 11 isoprene pyrophosphate synthetases (Undecaprenyl Pyrophosphate Synthetase)
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; Have the isolating polynucleotide of the full-length polypeptide of sequence described in SEQ ID NO:14 with the isolating polynucleotide of coding line plastochondria transit peptides and coding; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
D. the transcriptional of inferring of fatty acid metabolism
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; With the isolating polynucleotide of coding line plastochondria transit peptides and coding isolating polynucleotide as the full-length polypeptide of the transcriptional of inferring of fatty acid metabolism; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.The transcriptional of inferring of gene B1187 (SEQ ID NO:15) coding fatty acid metabolism.Transcriptional is partly characterized by their type of DNA binding domains and related factors.GntR type HTHDNA binding domains has partly characterized the classification by the fatty acid metabolism transcriptional of B1187 albumen (SEQ ID NO:16) example.
The transgenic plant of this embodiment can comprise any polynucleotide of the transcriptional of inferring of the fatty acid metabolism of encoding.Preferably, the transgenic plant of this embodiment comprise the polynucleotide of the full-length polypeptide of encoding, and wherein said polypeptide comprises gntR type HTH DNA binding domains.Preferably, this polynucleotide encoding comprise the transcription regulatory protein of the fatty acid metabolism polypeptide of gntR type HTH DNA binding domains, wherein said structural domain has the sequence of being made up of the 34th to the 53rd amino acids of SEQ ID NO:16.More preferably, this polynucleotide encoding comprise the transcription regulatory protein of the fatty acid metabolism polypeptide of transcription regulatory protein structural domain, wherein said transcription regulatory protein structural domain is made up of the 3rd to the 90th amino acids of SEQ ID NO:16.Most preferably, this polynucleotide encoding comprise the transcription regulatory protein of inferring of the fatty acid metabolism polypeptide of SEQ ID NO:4 the 1st to the 239th amino acids.
E. the nucleotide binding protein that has the P of G3E family ring structure territory
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; With the isolating polynucleotide of coding full-length polypeptide, described full-length polypeptide is the polypeptide that contains the Nucleotide binding domains; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.Gene B2173 (SEQ ID NO:17) coding contains the polypeptide (SEQ ID NO:18) in the P of G3E family ring GTP enzymatic structure territory.The P of G3E family ring GTP enzymatic structure territory is a feature there to be two distinctiveness motifs (near this mature polypeptide aminoterminal Walker A motif and a GTP specificity motif) partly.Walker A motif is G-x-x-x-x-G-K-S/T (SEQ ID NO:99).The function of Walker A motif is the triphosphoric acid part of settling bonded Nucleotide.GTP specificity motif is that the amino acid section of N/T-K-x-D (SEQ ID NO:100) and being considered to is better than other bases for guanine specificity is essential.Motif illustration in the protein described in Fig. 1 that this type of is conservative.
The transgenic plant of this embodiment can comprise the nucleotide binding protein that coding has the P of G3E family ring GTP enzymatic structure territory.Preferably, the transgenic plant of this embodiment have comprised coding and have had the polynucleotide of Nucleotide in conjunction with active full-length polypeptide, wherein this polypeptide comprises structural domain, described structural domain comprises the Walker A motif with the combination of GTP specificity motif, wherein Walker A motif has and is selected from the 9th to the 16th amino acids by SEQ ID NO:18, sequence in the group that SEQ ID NO:20 the 36th to the 43rd amino acids is formed, and GTP specificity motif has and is selected from the 152nd to the 155th amino acids by SEQ ID NO:18, sequence in the group that SEQ ID NO:20 the 191st to the 191st amino acids is formed.More preferably, this polynucleotide encoding has Nucleotide in conjunction with active full-length polypeptide, and wherein this polypeptide comprises the structural domain that is selected from the group of being made up of SEQ ID NO:18 the 6th to the 320th amino acids, SEQ ID NO:20 the 33rd to the 355th amino acids.Most preferably, the transgenic plant of this embodiment have comprised the protein-bonded polynucleotide of coding nucleotide, and wherein said nucleotide binding protein comprises SEQ ID NO:18 the 1st to the 328th amino acids; SEQ ID NO:20 the 1st to the 365th amino acids.
The membranin that F infers
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; Have the isolating polynucleotide of the membrane polypeptides that the total length of sequence described in SEQ ID NO:22 infers with coding; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.The membranin (SEQ ID NO:22) that gene B2670 (SEQ ID NO:21) coding is inferred.The transgenic plant of this embodiment can comprise any polynucleotide of the coding membranin of inferring, and wherein said membranin of inferring has the sequence that comprises SEQ ID NO:22 the 1st to 149 amino acids.
G peroxysome-coenzyme A synthetic enzyme
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; With the isolating polynucleotide of coding line plastochondria transit peptides and the isolating polynucleotide of coding total length peroxysome-coenzyme A synthetic enzyme polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.Gene YBR222C (the SEQ ID NO:23) peroxysome-coenzyme A synthetase albumen (SEQ ID NO:24) of encoding.Peroxysome-coenzyme A synthetic enzyme is a feature there to be the AMP binding domains with distinctive feature sequence partly.Characteristic sequence illustration in the peroxysome described in Fig. 2-coenzyme A synthetase albumen that this type of is conservative.
The transgenic plant of this embodiment can comprise any polynucleotide of coding peroxysome-coenzyme A synthetic enzyme.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has the full-length polypeptide of peroxysome-coenzyme A synthase activity, and wherein this polypeptide comprises to have and is selected from the 194th to the 205th amino acids by SEQ ID NO:24, SEQ ID NO:26 the 202nd to the 213rd amino acids, SEQ ID NO:28 the 214th to the 225th amino acids, SEQ ID NO:30 the 195th to the 206th amino acids, SEQ ID NO:32 the 175th to the 186th amino acids, SEQ ID NO:34 the 171st to the 182nd amino acids, SEQ ID NO:36 the 189th to the 200th amino acids, the AMP binding domains of the sequence in the group that SEQ ID NO:38 the 201st to the 212nd amino acids is formed.More preferably, this polynucleotide encoding has the full-length polypeptide of peroxysome-coenzyme A synthase activity, and wherein this polypeptide comprises and is selected from the 198th to the 456th amino acids by SEQ ID NO:24, SEQ ID NO:26 the 206th to the 477th amino acids, SEQ ID NO:28 the 218th to the 487th amino acids, SEQ ID NO:30 the 199th to the 468th amino acids, SEQ ID NO:32 the 179th to the 457th amino acids, SEQ ID NO:34 the 175th to the 452nd amino acids, SEQ ID NO:36 the 193rd to the 463rd amino acids, structural domain in the group that SEQ ID NO:38 the 205th to the 476th amino acids is formed.Most preferably, the transgenic plant of this embodiment comprise the polynucleotide of coding peroxysome-coenzyme A synthetic enzyme, and wherein said peroxysome-coenzyme A synthetic enzyme comprises the 1st to the 543rd amino acids of SEQ ID NO:24, the the 1st to the 569th amino acids of SEQ ID NO:26, the the 1st to the 565th amino acids of SEQ ID NO:28, the the 1st to the 551st amino acids of SEQ ID NO:30, the the 1st to the 560th amino acids of SEQ ID NO:32, the the 1st to the 543rd amino acids of SEQ ID NO:34, the the 1st to the 553rd amino acids of SEQ ID NO:36, the the 1st to the 568th amino acids of SEQ ID NO:38.
H. histone H 4 albumen
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; With the isolating polynucleotide of coding line plastochondria transit peptides and the isolating polynucleotide of coding total length histone H 4 polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.Gene YNL030W (SEQ ID NO:39) the histone H 4 albumen (SEQ ID NO:40) of encoding.Though found histone-like protein, histone is not present in the plastosome natively.Together with other nucleohistone, the H4 histone forms with nucleosome form (chromatinic primary structure unit) and is enclosed in karyon DNA octameric histone on every side.Histone H 4 protein part ground is feature there to be the distinctive feature sequence G-A-K-R-H (SEQ ID NO:101) that has between this protein the 14th and the 18th position.This conservative feature motif illustration in the histone H 4 albumen described in Fig. 3.
The transgenic plant of this embodiment can comprise the proteic any polynucleotide of coding histone H 4.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has the full-length polypeptide of histone H 4 albumen synthase activity, wherein this polypeptide comprises the structural domain that contains the histone H 4 label, and described histone H 4 label has the sequence that is selected from the group of being made up of SEQ ID NO:40 the 15th to the 19th amino acids, SEQ ID NO:56 the 15th to the 19th amino acids, SEQ ID NO:42 the 15th to the 19th amino acids, SEQ ID NO:44 the 15th to the 19th amino acids.More preferably, this polynucleotide encoding has the full-length polypeptide of histone H 4 protein-active, and wherein this polypeptide comprises the structural domain that is selected from the group of being made up of SEQ ID NO:40 the 3rd to the 92nd amino acids, SEQ ID NO:56 the 3rd to the 92nd amino acids, SEQ ID NO:42 the 3rd to the 92nd amino acids, SEQ ID NO:44 the 3rd to the 92nd amino acids.Most preferably, the transgenic plant of this embodiment comprise the polynucleotide of the histone H 4 of encoding, and wherein said histone H 4 comprises SEQ IDNO:40 the 1st to the 103rd amino acids, SEQ ID NO:56 the 1st to the 103rd amino acids, SEQ ID NO:42 the 1st to the 106th amino acids, SEQ ID NO:44 the 1st to the 105th amino acids.
I.SYM1 type conformity membrane albumen
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of coding chloroplast transit peptides and the proteic polynucleotide of coding total length SYM1 type conformity membrane; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
Gene YLR251W (SEQ ID NO:61) is SYM1 (" coercing derivable yeast Mpv17 ").Sym1 is the conformity membrane albumen that has vital role during the heat-shocked in the film transhipment.Hereinafter embodiment 2 shows that gene YLR251W (SEQ ID NO:61) are under USP promotor or the control of PCUbi promotor and under the growth conditions that is expressed in the water restriction of target chloroplast(id) or produce bigger plant when fully watering.Fig. 4 shows the comparison result of the representative SYM1 type polypeptide that this embodiment according to the present invention is used.
The transgenic plant of this embodiment can comprise any polynucleotide of coding SYM1 type conformity membrane polypeptide.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide of coding total length SYM1 type conformity membrane polypeptide, and wherein this polypeptide comprises and is selected from the 31st to the 171st amino acids by SEQ ID NO:62; SEQ ID NO:64 the 132nd to the 263rd amino acids; SEQ ID NO:50 the 131st to the 262nd amino acids; SEQ ID NO:52 the 12nd to the 145th amino acids; SEQ ID NO:54 the 134th to the 265th amino acids; Structural domain in the group of forming with SEQ ID NO:56 the 139th to the 272nd amino acids.Most preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has the SYM1 type conformity membrane polypeptide of following sequence, and wherein said sequence comprises the 1st to the 197th amino acids of SEQ ID NO:62; The the 1st to the 278th amino acids of SEQ ID NO:64; The the 1st to the 277th amino acids of SEQ ID NO:50; The the 1st to the 161st amino acids of SEQ ID NO:52; The the 1st to the 280th amino acids of SEQ ID NO:54; Or the 1st to the 293rd amino acids of SEQ ID NO:56.
J. vacuole pump subunit H polypeptide
In another embodiment, the invention provides and use expression cassette transgenic plant transformed, described expression cassette to comprise the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of the isolating polynucleotide of coding line plastochondria transit peptides and coding total length vacuolar proton pump subunit H polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.Gene YPR036W (SEQ ID NO:57) coding V-type ATP enzyme subunit H, it is a V-type atpase activity but non-its assembles necessary adjusting subunit in the yeast.Hereinafter embodiment 2 shows that gene YPR036W (SEQ ID NO:73) are producing bigger plant under the control of USP promotor and under the mitochondrial growth conditions that is expressed in the water restriction of target.Fig. 5 shows the comparison result of the representative V-type ATP enzyme subunit H polypeptide that this embodiment according to the present invention is used.
The transgenic plant of this embodiment can comprise any polynucleotide of coding V-type ATP enzyme subunit H polypeptide.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has V-type ATP enzyme subunit H active full-length polypeptide, wherein this polypeptide comprises structural domain, and this structural domain has the sequence that is selected from the group of being made up of SEQ ID NO:58 the 38th to the 470th amino acids, SEQ ID NO:60 the 19th to the 436th amino acids.Most preferably, this polynucleotide encoding comprises SEQ ID NO:58 the 1st to the 478th amino acids; The V-type ATP enzyme subunit H polypeptide of SEQ ID NO:60 the 1st to the 450th amino acids.
K.F-ATP enzyme subunit α polypeptide
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of the isolating polynucleotide of coding line plastochondria transit peptides and coding F-ATP enzyme subunit α polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.Gene SLL1326 (SEQ ID NO:61) coding is as the F-ATP enzyme subunit α of the essential component of F-ATP holoenzyme.Hereinafter embodiment 2 shows that gene SLL1326 (SEQ ID NO:61) are producing bigger plant under the control of ubiquitin promotor and under the mitochondrial growth conditions that is expressed in the water restriction of target.
The F-ATP enzyme is the original producer of ATP, and it uses the proton gradient by photosynthesis produced in oxidative phosphorylation in the plastosome or the chloroplast(id).The α of F-ATP enzyme and β subunit comprise with have sequence " P-[SAP]-[LIV]-[DNH]-{ the distinctive feature sequence of LKGN}-{F}-{S}-S-{DCPH}-S is the atp synthase structural domain of feature; wherein the amino acid position of square brackets inside can be described arbitrarily residue; the amino acid position of braces inside can be the arbitrary amino acid residue except listed, and the amino acid position with bracket only can not be that particular amino acid residue.Characteristic sequence illustration in the F-ATP enzyme subunit α albumen described in Fig. 6 that this type of is conservative.
The transgenic plant of this embodiment can comprise any polynucleotide of coding F-ATP enzyme subunit α.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has the full-length polypeptide of F-ATP enzyme subunit alpha active, wherein this polypeptide comprises structural domain, and this structural domain comprises the atp synthase characteristic sequence that is selected from the group of being made up of SEQ ID NO:62 the 356th to the 365th amino acids, SEQ ID NO:64 the 254th to the 263rd amino acids.More preferably, this polynucleotide encoding has the full-length polypeptide of F-ATP enzyme subunit alpha active, and wherein this polypeptide comprises the structural domain that is selected from the group of being made up of SEQ ID NO:62 the 149th to the 365th amino acids, SEQ ID NO:64 the 41st to the 263rd amino acids.Most preferably, the transgenic plant of this embodiment have comprised the polynucleotide of coding F-ATP enzyme subunit α, and wherein said F-ATP enzyme subunit α comprises SEQ ID NO:62 the 1st to the 503rd amino acids; SEQ ID NO:64 the 1st to the 388th amino acids.
L.F-ATP enzyme subunit beta polypeptides
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of the isolating polynucleotide of coding line plastochondria transit peptides and coding total length F-ATP enzyme subunit beta polypeptides; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.Gene SLR1329 (SEQ ID NO:65) coding is as the F-ATP enzyme subunit β of the essential component of F-ATP holoenzyme, as the α subunit.Hereinafter embodiment 2 shows that gene SLR1329 (SEQ ID NO:65) are producing bigger plant under the control of ubiquitin promotor and under the mitochondrial growth conditions that is expressed in the water restriction of target.F-ATP enzyme subunit β enzyme also partly with exist as to the described atp synthase characteristic sequence of α subunit " P-[SAP]-[LIV]-[DNH]-LKGN}-{F}-{S}-S-{DCPH}-S " be feature.Motif illustration in the F-ATP enzyme subunit β albumen described in Fig. 6 that this type of is conservative.
The transgenic plant of this embodiment can comprise any polynucleotide of coding F-ATP enzyme subunit β.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has the full-length polypeptide of F-ATP enzyme subunit 'beta ' activity, and wherein this polypeptide comprises the F-ATP enzyme subunit β that comprises SEQ ID NO:66 the 1st to the 483rd amino acids by polynucleotide encoding.
The M.ABC translocator
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of the isolating polynucleotide of coding line plastochondria transit peptides and coding total length abc transport protein polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.Gene SLR0977 (SEQ ID NO:67) coding abc transport albumen, it is to utilize the energy of ATP hydrolysis to come the transmembrane protein of transmembrane transport wide range of types substrate.Hereinafter embodiment 2 shows that gene SLR0977 (SEQ ID NO:67) are producing bigger plant under the control of ubiquitin promotor and under the mitochondrial growth conditions that is expressed in the water restriction of target.
The transgenic plant of this embodiment can comprise the proteic any polynucleotide of coding abc transport.Most preferably, the transgenic plant of this embodiment comprise the proteic polynucleotide of coding abc transport, and wherein said abc transport albumen comprises the 1st to 276 amino acids of SEQ ID NO:68.
N.PS-I subunit psaK polypeptide
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of the isolating polynucleotide of coding chloroplast transit peptides and coding total length PS-I subunit psaK polypeptide, wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.As shown in hereinafter embodiment 2, when comparing with the contrast arabidopsis thaliana, the transgenic arabidopsis plant that contains the cytoalgae gene ssr0390 (SEQ ID NO:69) of target chloroplast(id) shows the biomass that increases.The psaK subunit of ssr0390 genes encoding PS-I, described psaK subunit are feature there to be the distinctiveness PsaGK characteristic sequence of representing the psaG/psaK family gene partly.Photosystem I psaGK characteristic sequence is [GTND]-[FPMI]-x-[LIVMH]-x-[DEAT]-x (2)-[GA]-x-[GTAM]-[STA]-x-G-H-x-[LIVM]-[GAS], wherein the amino acid position of square brackets inside can be described arbitrarily residue.Protein psaK is the hydrophobicity small protein with two membrane spaning domains (14th to 34th amino acids of SEQ ID NO:70 and 61st to 81st amino acids) relevant with psaG in the plant.The psaGK characteristic sequence is present in the 56th to the 73rd residue position and thereby almost completely is positioned at the second membrane spaning domain inside.
The transgenic plant of this embodiment can comprise any polynucleotide of coding total length psaK subunit.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has psaK active full-length polypeptide, wherein this polypeptide comprises the PSI_PsaK label, and described PSI_PsaK label comprises the amino acid of SEQ ID NO:2 the 14th to the 86th amino acids.More preferably, the transgenic plant of this embodiment comprise the polynucleotide of the I of encoded light system reactive center psaK subunit, and wherein said photosystem I reactive center psaK subunit has the sequence that comprises SEQ ID NO:2 the 1st to 86 amino acids.
O. ferredoxin
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of the isolating polynucleotide of coding line plastochondria transit peptides and coding total length ferredoxin polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.As shown in hereinafter embodiment 2, when comparing with the contrast arabidopsis thaliana, the mitochondrial transgenic arabidopsis plant of target that contains cytoalgae gene sll1382 (SEQ ID NO:71) shows the biomass that increases.The sll1382 genes encoding be the ferredoxin (PetF) of feature partly there to be the Fer2 characteristic sequence.This type of characteristic sequence is illustration in the ferredoxin described in Fig. 7.
The transgenic plant of this embodiment can comprise any polynucleotide of the ferredoxin of encoding.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has the active full-length polypeptide of ferredoxin, and wherein this polypeptide comprises and is selected from the 11st to the 87th amino acids by SEQ ID NO:72; SEQ ID NO:74 the 12nd to the 88th amino acids; Fer2 characteristic sequence in the group that SEQ ID NO:76 the 63rd to the 139th amino acids is formed.More preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has the ferredoxin polypeptide of following sequence, and wherein said sequence comprises the 1st to the 122nd amino acids of SEQ ID NO:72; The the 1st to the 128th amino acids of SEQ ID NO:74; The the 1st to the 179th amino acids of SEQ ID NO:76.
P. flavodoxin
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of coding chloroplast transit peptides; Comprise the isolating polynucleotide of the total length flavodoxin polypeptide of SEQ ID NO:78 the 6th to the 160th amino acids with coding; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.As shown in hereinafter embodiment 2, when comparing with the contrast arabidopsis thaliana, the transgenic arabidopsis plant that contains the target chloroplast(id) of cytoalgae gene sll0248 (SEQ ID NO:77) shows the biomass that increases.Sll0248 genes encoding flavodoxin and be feature partly there to be flavodoxin _ 1 characteristic sequence as SEQ ID NO:78 the 6th to the 160th amino acids representative.
The transgenic plant of this embodiment can comprise any polynucleotide of coding total length flavodoxin polypeptide, and wherein said total length flavodoxin polypeptide comprises the 6th to 160 amino acids of SEQ ID NO:78.Preferably, the transgenic plant of this embodiment comprise the polynucleotide of coding total length flavodoxin, and wherein said total length flavodoxin comprises the 1st to 170 amino acids of SEQ ID NO:1.
Q.PS-I psaF polypeptide
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of the isolating polynucleotide of coding chloroplast transit peptides and coding total length PS-I psaF polypeptide, wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.As shown in hereinafter embodiment 2, when comparing with the contrast arabidopsis thaliana, the transgenic arabidopsis plant that contains the target chloroplast(id) of cytoalgae gene sll0819 (SEQ ID NO:79) shows the biomass that increases.The sll0819 genes encoding be the PS-I subunit III (PsaF) of feature partly there to be the PSI_PsaF characteristic sequence.This type of characteristic sequence is illustration in the PS-I subunit III albumen described in Fig. 8.
The transgenic plant of this embodiment can comprise any polynucleotide of coding PS-I subunit III.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has the active full-length polypeptide of PS-I subunit III, and wherein this polypeptide comprises and is selected from the 3rd to the 158th amino acids by SEQ ID NO:80; SEQ ID NO:82 the 43rd to the 217th amino acids; SEQ ID NO:84 the 46th to the 220th amino acids; SEQ ID NO:86 the 50th to the 224th amino acids; PSI_PsaF characteristic sequence in the group of forming with SEQ ID NO:88 the 50th to the 224th amino acids.More preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has the plant PS-I subunit III of following sequence, and wherein said sequence comprises the 1st to the 217th amino acids of SEQ ID NO:82; The the 1st to the 220th amino acids of SEQ ID NO:84; The the 1st to the 224th amino acids of SEQ ID NO:86; Or the 1st to the 224th amino acids of SEQ ID NO:88.
R. cytochrome c 553 albumen
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of the isolating polynucleotide of coding line plastochondria transit peptides and coding total length cytochrome c 553 albumen (petJ) polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the biomass that increases.As shown in hereinafter embodiment 2, when comparing with the contrast arabidopsis thaliana, the mitochondrial transgenic arabidopsis plant of target that contains cytoalgae gene sll1796 (SEQ ID NO:89) shows the output that increases.
Gene sll1796 (SEQ ID NO:89) Codocyte pigment C553.Cytochrome C 553 (RetJ) is also referred to as cytochrome c 6, participates in the electron transport of photosynthetic property.PetJ plays a role as the electron carrier between film mating type cytochrome b 6-f and the photosystem I, and this is a function of being fulfiled by plastocyanin in higher plant.Photosynthetic property electron transport from cytochrome b f complex body to PS-I can be by cytochrome c 6 or plastocyanin mediation, and this depends on the concentration of copper in the growth medium.Cytochrome c 553 protein part ground are feature there to be the cytopigment _ C characteristic sequence as SEQ ID NO:90 the 38th to the 116th amino acids representative.The transgenic plant of this embodiment can comprise the proteic any polynucleotide of Codocyte pigment c553.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has cytochrome c 553 active full-length polypeptides, wherein this polypeptide comprises cytopigment _ C characteristic sequence, and described cytopigment _ C characteristic sequence comprises SEQ ID NO:90 the 38th to the 116th amino acids.More preferably, the transgenic plant of this embodiment comprise the polynucleotide of Codocyte pigment c553 polypeptide, and wherein said cytochrome c 553 polypeptide have the sequence that comprises SEQ ID NO:90 the 1st to 120 amino acids.
S.PS_II W polypeptide
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of the isolating polynucleotide of coding line plastochondria transit peptides and coding total length PS-II W (PsbW) polypeptide, wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.As shown in hereinafter embodiment 2, when comparing with the contrast arabidopsis thaliana, the mitochondrial transgenic arabidopsis plant of target that contains cytoalgae gene sll1739 (SEQ ID NO:91) shows the biomass that increases.It is the psbW of feature there to be the PsbW characteristic sequence as SEQ ID NO:92 the 5th to the 120th amino acids representative partly that gene slr1739 has encoded.
The transgenic plant of this embodiment have comprised the proteic any polynucleotide of total length PsbW that coding comprises the PsbW characteristic sequence, and wherein said PsbW characteristic sequence comprises the 5th to 120 amino acids of SEQ ID NO:92.More preferably, the transgenic plant of this embodiment comprise the active polynucleotide of coding PsbW, and wherein said PsbW activity has the sequence that comprises SEQ ID NO:92 the 1st to 122 amino acids.
T. uroporphyrin-III C-methyltransgerase
In another embodiment, the invention provides the isolating polynucleotide of using expression cassette transgenic plant transformed, described expression cassette to comprise the coding promotor that is in effective connection; The isolating polynucleotide of the isolating polynucleotide of coding chloroplast transit peptides and coding total length uroporphyrin-III C-methyltransgerase (CobA) polypeptide; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the biomass that increases.As shown in hereinafter embodiment 2, when comparing with the contrast arabidopsis thaliana, the transgenic arabidopsis plant that contains the target chloroplast(id) of cytoalgae gene sll0378 (SEQ ID NO:93) shows the output that increases.Gene sll0378 (the SEQ ID NO:93) uroporphyrin-III C-methyltransgerase (CobA) of encoding.Uroporphyrin-III c-methyltransgerase is a feature there to be TP_ methylase characteristic sequence partly.
The transgenic plant of this embodiment can comprise the polynucleotide of plant arbitrarily of coding uroporphyrin-III c-methyltransgerase.Preferably, the transgenic plant of this embodiment comprise the polynucleotide that coding has the full-length polypeptide of uroporphyrin-III c-methyl transferase activity, and wherein said full-length polypeptide has the sequence that comprises SEQ ID NO:94 the 1st to 263 amino acids.
U. preceding corrin-6b methylase
In another embodiment, the invention provides and use the expression cassette transgenic plant transformed, described expression cassette comprises the isolating polynucleotide of the coding promotor that is in effective connection and the isolating polynucleotide of the preceding corrin of coding total length-6b methylase polypeptide, and wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.The expression cassette of this embodiment can randomly comprise the isolating polynucleotide of coding line plastochondria transit peptides.As shown in hereinafter embodiment 2, when comparing with the contrast arabidopsis thaliana, the transgenic arabidopsis plant that contains cytoalgae gene slr1368 (SEQ ID NO:95) shows the biomass that increases.It is the preceding corrin-6b methylase of feature there to be methyltransgerase _ 12 characteristic sequences partly that gene slr1368 has encoded.
The transgenic plant of this embodiment can comprise any polynucleotide of the preceding corrin of coding-6b methylase.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has preceding corrin-active full-length polypeptide of 6b methylase, wherein this polypeptide comprises methyltransgerase _ 12 characteristic sequences, and described methyltransgerase _ 12 characteristic sequences comprise SEQ ID NO:96 the 45th to the 138th amino acids.Most preferably, the transgenic plant of this embodiment comprise the polynucleotide of the preceding corrin of coding-6b methylase, and wherein said preceding corrin-6b methylase has the sequence that comprises SEQ ID NO:96 the 1st to 197 amino acids.
Corrin before the V decarboxylation-6y methylase
In another embodiment, the invention provides and use the expression cassette transgenic plant transformed, described expression cassette comprises isolating polynucleotide and the preceding corrin of coding total length decarboxylation-6y c5 of the coding promotor that is in effective connection, the isolating polynucleotide of 15-Methyl transporters enzyme polypeptide, wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.The expression cassette of this embodiment can randomly comprise the isolating polynucleotide of coding line plastochondria transit peptides.As shown in hereinafter embodiment 2, with contrast arabidopsis thaliana when comparing, contain the target of cytoalgae gene sll0099 (SEQ ID NO:97) and not the mitochondrial transgenic arabidopsis plant of target show the biomass of increase.It is corrin-6y methylase activity before the decarboxylation of feature that gene sll0099 has encoded partly there to be TP_ methylase characteristic sequence.
The transgenic plant of this embodiment can comprise the preceding corrin of coding decarboxylation-active any polynucleotide of 6y methylase.Preferably, the transgenic plant of this embodiment have comprised the polynucleotide that coding has corrin-6y methylase active full-length polypeptide before the decarboxylation, wherein this polypeptide comprises TP_ methylase characteristic sequence, and described TP_ methylase characteristic sequence comprises SEQ ID NO:98 the 1st to the 195th amino acids.Most preferably, the transgenic plant of this embodiment comprise the polynucleotide of the preceding corrin of coding decarboxylation-6y methylase, and corrin before the wherein said decarboxylation-6y methylase has the sequence that comprises SEQ ID NO:98 the 1st to 425 amino acids.
The present invention also provides the seed for isozygotying of described expression cassette (being also referred to as " transgenosis " herein) herein, shows the output that increases when wherein the transgenic plant that go out from described cultivating seeds are compared with the wild-type kind of this plant.The present invention also provide following by or the product that from the transgenic plant of expressing described polynucleotide, their plant part and its seed, produces.This product can use several different methods well known in the art to obtain.As used herein, vocabulary " product " includes but not limited to food, feed, food fill-in, feed supplement, fiber, makeup or medicine.Food is considered as the composition that is used for nutrition or is used to supplement the nutrients.Especially animal-feed and animal-feed fill-in are considered as food.The present invention also provides the agricultural-food that any one produced by described transgenic plant, plant part and plant seed.Agricultural-food include but not limited to plant milk extract, protein, amino acid, carbohydrate, fat, oils, polymkeric substance, VITAMIN etc.
The present invention also provides isolating polynucleotide, and it has and is selected from the NO:19 by SEQ ID, SEQ ID NO:25, SEQ ID NO:27, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:33, SEQ ID NO:37, SEQ ID NO:41, SEQ ID NO:43, SEQ ID NO:63, SEQ ID NO:49, SEQ ID NO:51, SEQ ID NO:53, SEQ ID NO:59, SEQ ID NO:63, SEQ ID NO:73, SEQ ID NO:75, SEQ ID NO:81, SEQ ID NO:83, sequence in the group that SEQ ID NO:85 and SEQ ID NO:87 form.Isolating polynucleotide of the present invention also comprise the isolating polynucleotide of coded polypeptide, and described polypeptide has and is selected from the NO:20 by SEQ ID, SEQ ID NO:26, SEQ ID NO:28, SEQ ID NO:30, SEQ ID NO:32, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:44, SEQ ID NO:64, SEQ ID NO:50, SEQ ID NO:52, SEQ ID NO:54, SEQ ID NO:60, SEQ ID NO:64, SEQ ID NO:74, SEQ ID NO:76, SEQ ID NO:82, SEQ ID NO:84, aminoacid sequence in the group that SEQ ID NO:86 and SEQ ID NO:88 form.The sequence information that can use standard molecular biological technique (for example to use automatic dna synthesizer) with is herein provided separates polynucleotide of the present invention.
Isolating polynucleotide of the present invention comprise the homologue of table 1 polynucleotide." homologue " is defined as two kinds of nucleic acid or the polypeptide that has similar or substantially the same nucleotide sequence or aminoacid sequence respectively in this article.Homologue comprises as the allele variant, analogue of hereinafter definition with directly to homologue.As used herein, term " analogue " refers to have same or analogous function, but two kinds of nucleic acid of in incoherent biology, independently evolving.As used herein, term " directly to homologue " refers to from different plant species, but forms two kinds of nucleic acid from common my late grandfather's gene evolution by species.Term " homologue " also comprises because of the genetic code degeneracy and is different from one of nucleotide sequence shown in the table 1 and the nucleic acid molecule of the phase homopolypeptide of therefore encoding.
For (for example determining two seed amino acid sequences, one of peptide sequence of table 1 and homologue thereof) sequence identity percentage ratio, compare described sequence (for example, can in the sequence of a peptide species, import the room) for the best comparison purpose with the comparison of optimization with another kind of polypeptide or nucleic acid.The amino-acid residue at more corresponding subsequently amino acid position place.When the position in the sequence occupied as the same amino acid residue of correspondence position in by another sequence, then described molecule was identical in this position.The comparison of same type can be carried out between two kinds of nucleotide sequences.
Preferably, the isolating amino acid homology thing of polypeptide of the present invention, analogue and the complete amino acid sequence directly identified in homologue and table 1 be at least about 50-60%, preferably at least about 60-70% with more preferably at least about 70-75%, 75-80%, 80-85%, 85-90% or 90-95%, and most preferably at least about 96%, 97%, 98%, 99% or more how same.In another preferred embodiment, isolating nucleic acid homologue of the present invention comprise with nucleotide sequence shown in the table 1 at least about 40-60%, preferably at least about 60-70%, more preferably at least about 70-75%, 75-80%, 80-85%, 85-90% or 90-95%, and even more preferably at least about 95%, 96%, 97%, 98%, 99% or how same nucleotide sequence.
For the purposes of the present invention, use Align 2.0 (Myers and Miller, CABIOS (1989) 4:11-17) at whole parameter settings to the situation of default configuration or Vector NTI 9.0 (PC) software package (Invitrogen, 1600 Faraday Ave., Carlsbad CA92008) determines sequence identity percentage ratio between two kinds of nucleic acid or the peptide sequence.For the identity percentage ratio that calculates with Vector NTI, the identity percentage ratio that point penalty 6.66 is used for determining two kinds of nucleic acid is extended in room opening point penalty 15 and room.The identity percentage ratio that point penalty 0.1 is used for determining two peptide species is extended in room opening point penalty 10 and room.All other parameter is set under default configuration., adopt under the situation of blosum62 matrix purpose (Clustal W algorithm) for multiple ratio, room opening point penalty is 10 and to extend point penalty be 0.05 in the room.Be to be understood that for the purpose of determining sequence identity when dna sequence dna and the comparison of RNA sequence, thymidylic acid is equal to uridylate.
With the homologue of listed polypeptide in the table 1, analogue and directly can be based on the identity of they and described polypeptide to the corresponding nucleic acid molecule of homologue, use the polynucleotide of coding corresponding polypeptide or based on the primer of these polynucleotide as hybridization probe, according to the standard hybridization technique, under stringent hybridization condition, separate.As herein with regard to DNA to the hybridization aspect of southern blotting technique thing used with regard to, term " stringent condition " refers to hybridize in 10 * Denhart solution, 6 * SSC, 0.5%SDS and 100 μ g/ml sex change salmon sperm DNAs at 60 ℃ and spends the night.The trace thing 62 ℃ successively at 3 * SSC/0.1%SDS, subsequently at 1 * SSC/0.1%SDS and in 0.1 * SSC/0.1%SDS, wash each 30 minutes at last.Also as used herein, in preferred embodiments, phrase " stringent condition " refers in 6 * SSC solution 65 ℃ of hybridization.In another embodiment, " height stringent condition " refers to hybridize in 10XDenhart solution, 6 * SSC, 0.5%SDS and 100 μ g/ml sex change salmon sperm DNAs at 65 ℃ and spends the night.The trace thing 65 ℃ successively at 3 * SSC/0.1%SDS, subsequently at 1 * SSC/0.1%SDS and in 0.1 * SSC/0.1%SDS, wash each 30 minutes at last.The method that is used to carry out nucleic acid hybridization is well known in the art.
Isolating polynucleotide used among the present invention can be optimized, that is, genetically engineered to increase its expression in given plant or animal.The nucleic acid provide plant to optimize is provided, the dna sequence dna that can modify this gene with: 1) comprise the codon of the plant gene institute preference of highly being expressed; 2) comprise the A+T content that is in the nucleotide bases composition that exists in a large number in the plant; 3) form the plant homing sequence; Or 4) elimination causes RNA stabilization removal, improper polyadenylation, degraded and terminated sequence, or eliminates the sequence that can form secondary hairpin structure or RNA splice site; Or 5) eliminate the antisense open reading-frame (ORF).Can realize the expression that nucleic acid increases in plant by the distribution frequency of utilizing in the general plant or codon uses in the specified plant.Can be at EPA 0359472; EPA 0385962; PCT application number WO 91/16432; U.S. Patent number 5,380,831; U.S. Patent number 5,436,391; Perlack etc., 1991, Proc.Natl.Acad.Sci.USA 88:3324-3328 and Murray etc., 1989, find among the Nucleic Acids Res.17:477-498 to be used for the method that plant is optimized expression of nucleic acid.
The present invention also provides the recombinant expression vector that comprises expression cassette, wherein said expression cassette is selected from the group of being made up of following expression cassette: a) expression cassette, and it comprises separation polynucleotide and the coding that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf and has the NO:2 as SEQ ID; SEQ ID NO:4; SEQ ID NO:6; SEQ ID NO:8; Or the isolating polynucleotide of the full-length polypeptide of sequence described in the SEQ ID NO:22; B) expression cassette, it comprises the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of coding plastid transit peptides; Have the isolating polynucleotide of the full-length polypeptide of sequence described in SEQ ID NO:10, SEQ ID NO:12 or SEQ ID NO:14 with coding; C) expression cassette, it comprises the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of coding line plastochondria transit peptides; Have the isolating polynucleotide of total length transcriptional of the fatty acid metabolism of gntR type HTH DNA binding domains with coding, described gntR type HTH DNA binding domains comprises the 34th to the 53rd amino acids of SEQ ID NO:16; D) expression cassette, it comprises the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; Has G3E with coding, the isolating polynucleotide of the full-length polypeptide in P ring structure territory, described G3E, P ring structure territory has comprised Walker A motif with sequence described in SEQ ID NO:99 and the GTP specificity motif with sequence described in SEQ ID NO:100; E) expression cassette, it comprises the isolating polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of coding line plastochondria transit peptides; Comprise the isolating polynucleotide of the total length peroxysome-coenzyme A synthetic enzyme polypeptide of AMP binding domains with coding, described AMP binding domains is selected from the 194th to the 205th amino acids by SEQ ID NO:24, SEQ ID NO:26 the 202nd to the 213rd amino acids, SEQ ID NO:28 the 214th to the 225th amino acids, SEQ ID NO:30 the 195th to the 206th amino acids, SEQ ID NO:32 the 175th to the 186th amino acids, SEQ ID NO:34 the 171st to the 182nd amino acids, SEQ ID NO:36 the 189th to the 200th amino acids, the group that SEQ ID NO:38 the 201st to the 212nd amino acids is formed; F) expression cassette, it comprises the isolating polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of coding line plastochondria transit peptides; Have the isolating polynucleotide of the total length histone H 4 polypeptide of G-A-K-R-H (SEQ ID NO:101) characteristic sequence structural domain with coding, described G-A-K-R-H characteristic sequence structural domain is selected from the 3rd to the 92nd amino acids by SEQ ID NO:40; SEQ ID NO:56 the 3rd to the 92nd amino acids; The group that SEQ ID NO:42 the 3rd to the 92nd amino acids and SEQ ID NO:44 the 3rd to the 92nd amino acids are formed; G) expression cassette, it comprises the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf or the isolating polynucleotide of coding constitutive promoter; The isolating polynucleotide of coding chloroplast transit peptides; With the proteic polynucleotide of coding total length SYM1 type conformity membrane; H) expression cassette, it comprises the isolating polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of the isolating polynucleotide of coding line plastochondria transit peptides and coding total length vacuolar proton pump subunit H polypeptide; I) expression cassette, it comprises the separation polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of coding line plastochondria transit peptides; Comprise the isolating polynucleotide of the F-ATP enzyme subunit α polypeptide of atp synthase structural domain with coding, described atp synthase structural domain is selected from the 356th to the 365th amino acids by SEQ ID NO:62; The group that SEQ ID NO:64 the 254th to the 263rd amino acids is formed; J) expression cassette, it comprises the isolating polynucleotide that the coding that is in effective connection can strengthen the promotor of genetic expression in the leaf; The isolating polynucleotide of coding line plastochondria transit peptides; Have the isolating polynucleotide of the total length F-ATP enzyme subunit beta polypeptides of sequence described in SEQ ID NO:66 with coding; K) expression cassette, it comprises the isolating polynucleotide of the coding promotor that is in effective connection, the isolating polynucleotide of coding line plastochondria transit peptides; Have the isolating polynucleotide of the total length abc transport protein polypeptide of sequence described in SEQ ID NO:68 with coding; L) expression cassette, it comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding chloroplast transit peptides; Have the isolating polynucleotide of the total length PS-I subunit psaK polypeptide of psaGK label with coding, described psaGK label comprises the 56th to the 73rd amino acids of SEQ ID NO:70; M) expression cassette, it comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding line plastochondria transit peptides; Comprise the isolating polynucleotide of the total length ferredoxin polypeptide of Fer2 characteristic sequence with coding, described Fer2 characteristic sequence is selected from the 11st to the 87th amino acids by SEQ ID NO:72; SEQ ID NO:74 the 12nd to the 88th amino acids; The group that SEQ ID NO:76 the 63rd to the 139th amino acids is formed; N) expression cassette, it comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding chloroplast transit peptides; Have the isolating polynucleotide of the total length flavodoxin polypeptide of flavodoxin _ 1 characteristic sequence with coding, described flavodoxin _ 1 characteristic sequence comprises the 6th to the 160th amino acids of SEQ ID NO:78; O) expression cassette, it comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding chloroplast transit peptides; Comprise the isolating polynucleotide of the total length PS-IpsaF polypeptide of PSI_PsaF characteristic sequence with coding, described PSI_PsaF characteristic sequence is selected from the 3rd to the 158th amino acids by SEQ ID NO:80; SEQ ID NO:82 the 43rd to the 217th amino acids; SEQ ID NO:84 the 46th to the 220th amino acids; The group that SEQ ID NO:86 the 50th to the 224th amino acids and SEQ ID NO:88 the 50th to the 224th amino acids are formed; P) expression cassette, it comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding line plastochondria transit peptides; Isolating polynucleotide with coding line plastochondria transit peptides; Have the isolating polynucleotide of total length cytochrome c 553 (PetJ) polypeptide of PSI_PsaF characteristic sequence with coding, described PSI_PsaF characteristic sequence comprises the 38th to the 116th amino acids of SEQ ID NO:90; Q) expression cassette, it comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding line plastochondria transit peptides; Have the isolating polynucleotide of total length PS-II W (PsbW) polypeptide of cytochrome C characteristic sequence with coding, described cytochrome C characteristic sequence comprises the 5th to the 120th amino acids of SEQ ID NO:92; R) expression cassette, it comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding chloroplast transit peptides; Have the isolating polynucleotide of total length uroporphyrin-III c-methyltransgerase (CobA) polypeptide of sequence described in SEQ ID NO:92 with coding; S) expression cassette, it comprises the isolating polynucleotide of the isolating polynucleotide of the coding promotor that is in effective connection and the preceding corrin of total length-6b methylase that coding has methyltransgerase _ 12 characteristic sequences, and described methyltransgerase _ 12 characteristic sequences comprise the 45th to the 138th amino acids of SEQ ID NO:96; And t) expression cassette, it comprises the isolating polynucleotide of the coding promotor that is in effective connection and the preceding corrin of total length decarboxylation-6y c5 that coding has TP_ methylase characteristic sequence, the isolating polynucleotide of 15-methyltransgerase, described TP_ methylase characteristic sequence comprises the 1st to the 195th amino acids of SEQ ID NO:98.
In another embodiment, recombinant expression vector of the present invention comprises isolating polynucleotide, and it has and is selected from the NO:19 by SEQ ID; SEQ ID NO:25; SEQ ID NO:27; SEQ ID NO:29; SEQ ID NO:31; SEQ ID NO:33; [SEQ ID NO:35? ]; SEQ ID NO:37; SEQ ID NO:41; SEQ ID NO:43; SEQ ID NO:63; SEQ ID NO:49; SEQ ID NO:51; SEQ ID NO:53; [SEQ ID NO:55? ]; SEQ ID NO:59; SEQ ID NO:63; SEQ ID NO:73; SEQ ID NO:75; SEQ ID NO:81; SEQ ID NO:83; Sequence in the group that SEQ ID NO:85 and SEQ ID NO:87 form.In addition, recombinant expression vector of the present invention comprises the isolating polynucleotide of coded polypeptide, and described polypeptide has and is selected from the NO:20 by SEQ ID, SEQ ID NO:26, SEQ ID NO:28, SEQ ID NO:30, SEQ ID NO:32, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:44, SEQ ID NO:64, SEQ ID NO:50, SEQ ID NO:52, SEQ ID NO:54, SEQ ID NO:60, SEQ ID NO:64, SEQ ID NO:74, SEQ ID NO:76, SEQ ID NO:82, SEQ ID NO:84, aminoacid sequence in the group that SEQ ID NO:86 and SEQ ID NO:88 form.
Recombinant expression vector of the present invention comprises one or more adjusting sequences of selecting based on the host cell that is ready to use in expression, and described adjusting sequence effectively is connected with isolating polynucleotide to be expressed.As used with regard to the recombinant expression vector aspect herein, " being in effective connection " or " effectively connect " mean polynucleotide of interest with regulate sequence when being imported into host cell with this carrier (for example in bacillary or plant host cell) allow the mode of described polynucleotide expression to be connected.Term " adjusting sequence " is used to comprise promotor, enhanser and other expression controlling elements (for example, polyadenylation signal).
The one or more adjusting sequences that effectively are connected with described polynucleotide this is combined in the common element that this area is called " expression cassette ", and wherein said adjusting sequence is selected based on the host cell that is ready to use in expression.This expression cassette can also contain following defined chloroplast(id) or the mitochondrial transport sequence that is connected with described polynucleotide.Often expression cassette is described as " construct " in the art and uses these two terms herein with being equal to.
As mentioned above, certain embodiments of the present invention are used the promotor that can strengthen genetic expression in the leaf.In some embodiments, this promotor is the leaf specificity promoter.The leaf specificity promoter can be used for these embodiments of the present invention arbitrarily.These type of many promotors are known, for example, USP promotor (people (1991) Mol.Gen.Genet.225 such as Baeumlein from broad bean (Vicia faba), 459-67), photoinduction type gene such as ribulose-1,5-bisphosphate, the promotor of 5-bisphosphate carboxylase (rbcS promotor), the promotor (Cab) of the protein-bonded gene of coding chlorophyll a/b, the Rubisco activating enzymes, Arabidopis thaliana chloroplast(id) glyceraldehyde 3-phosphate dehydro-genase B-subunit (people such as Kwon. (1994) Plant Physiol.105,357-67) with other leaf specificity promoter such as Aleman, I. (2001) separation and sign are from the leaf specificity promoter (Isolation and characterization of leaf-specific promoters from alfalfa (Medicago sativa)) of clover, Master's thesis, Mexico State University, Los Cruces, those promotors of identifying among the NM.
In other embodiments of the present invention, use root or sprout specificity promoter.For example, super promotor all provides high level expression (people (1995) Plant such as Ni J.7:661-676) in root and sprout.Other root-specific promoter include but not limited to the TobRB7 promotor (people (1991) Plant Cell 3 such as Yamamoto, 371-382), the rolD promotor (people (1991) Plant Science 79 such as Leach, 69-76); CaMV 35S structural domain A promotor (people (1989) Science 244 such as Benfey, 174-181) etc.
In other embodiments, use constitutive promoter.Constitutive promoter is activated under most of conditions.The example that is applicable to the constitutive promoter in these embodiments comprises the parsley ubiquitin promotor described in the WO2003/102198 (SEQ ID NO:102), CaMV 19S and 35S promoter, sX CaMV 35S promoter, the Sep1 promotor, the rice actin promoter, the Arabidopis thaliana actin promoter, corn ubiquitin promotor, pEmu, radix scrophulariae (figwort) mosaic virus 35 S promoter, the Smas promotor, super promotor (U.S. Patent number 5,955,646), the GRP1-8 promotor, cinnamyl-alcohol dehydrogenase promotor (U.S. Patent number 5,683,439), from the promotor of agrobatcerium T-DNA (as mannopine synthase, the promotor of nopaline synthase and octopine synthase), carboxydismutase small subunit (ssuRUBISCO) promotor etc.
According to the present invention, chloroplast transit sequence refer to the to encode nucleotide sequence of chloroplast transit peptides.Chloroplast targeted sequence is known in the art and comprises ribulose-1,5-bisphosphate, the chloroplast(id) small subunit of 5-bisphosphate carboxylase (Rubisco) (people such as de Castro Silva Filho, (1996), Plant Mol.Biol.30:769-780; People such as Schnell, (1991), J.Biol.Chem.266 (5): 3335-3342); 5-(enolpyrul) shikimic acid-3-phosphate synthase (EPSPS) (people such as Archer, (1990), J.Bioenerg.Biomemb.22 (6): 789-810); Tryptophan synthetase (people such as Zhao, (1995), J.Biol.Chem.270 (11): 6081-6087); Plastocyanin (people such as Lawrence, (1997), J.Biol.Chem.272 (33): 20357-20363); Chorismate synthase (people such as Schmidt, (1993), J.Biol.Chem.268 (36): 27447-27457); Ferredoxin (people such as Jansen. (1988) Curr.Genetics13:517-522) (SEQ ID NO:111); Nitrite reductase (people (1988) MGG212:20-26 such as Back); With light harvesting chlorophyll a/b conjugated protein (LHBP) (people (1988) J.Biol.Chem.263:14996-14999 such as Lamppa).Also referring to people (1991) Plant Mol.Biol.Rep.9:104-126 such as Von Heijne; People such as Clark (1989) J.Biol.Chem.264:17544-17550; People such as Della-Cioppa (1987) Plant Physiol.84:965-968; People such as Romer (1993) Biochem.Biophys.Res.Commun.196:1414-1421; With people (1986) Science 233:478-481 such as Shah.
As definition herein, the mitochondrial transport sequence refers to that coding line plastochondria presequence and guides protein to mitochondrial nucleotide sequence.The example of plastosome presequence comprises the group of being made up of ATP enzyme subunit, atp synthase subunit, Rieske-FeS albumen, Hsp60, malate dehydrogenase (malic acid dehydrogenase), Oxalacetic transacetase, aconitase, isocitric enzyme, pyruvic oxidase, malic enzyme, glycine decarboxylase, serine hydroxymethylase, isovaleryl-CoA dehydrogenase and superoxide-dismutase.This type of transit peptides is known in the art.See, for example, people (1991) Plant Mol.Biol.Rep.9:104-126 such as Von Heijne; People such as Clark (1989) J.Biol.Chem.264:17544-17550; People such as Romer (1993) Biochem.Biophys.Res.Commun.196:1414-1421; People such as Faivre-Nitschke (2001) Eur J Biochem 268 1332-1339; Deng people (1999) 39:1275-1282 (SEQ ID NO:109 and SEQ ID NO:107); With people (1986) Science 233:478-481 such as Shah.
In a preferred embodiment of the invention, in from the vegetable cell (for example, spermatophyte is as crop plants) of higher plant, express listed polynucleotide in the table 1.Polynucleotide can be by any means " importing " vegetable cell, and described method comprises infection protocol, conversion or transduction method, electroporation, particle bombardment method, agroinfection etc.Disclose and be used to transform or the appropriate method of transfection of plant cells, for example used in U.S. Patent number 4,945,050; 5,036,006; 5,100,792; 5,302,523; 5,464,765; 5,120,657; The particle bombardment method of describing in 6,084,154 grades.More preferably, use as U.S. Patent number 5,591,616; 5,731,179; 5,981,840; 5,990,387; 6,162,965; 6,420,630, the Agrobacterium-mediated Transformation method of describing in U.S. Patent Application Publication No. 2002/0104132 grade can prepare transgenic corn seed of the present invention.Can use as european patent number 0424047, U.S. Patent number 5,322,783, european patent number 0397687, U.S. Patent number 5,376,543 or U.S. Patent number 5,169, each technology described in 770 is carried out the conversion of soybean.The specific examples that can in PCT application number WO93/07256, find wheat to transform.Can use in U.S. Patent number 5,004,863; 5,159,135; Disclosed method transforms cotton in 5,846,797 grades.Can use in U.S. Patent number 4,666,844; 5,350,688; 6,153,813; 6,333,449; 6,288,312; 6,365,807; Disclosed method transforms rice in 6,329,571 grades.Can for example use several different methods, as in U.S. Patent number 5,188,958; 5,463,174; 5,750,871; EP1566443; Disclose those methods among the WO02/00900 etc., transform the canola oil dish.Other methods for plant transformation is at for example U.S. Patent number 5,932,782; 6,153,811; 6,140,553; 5,969,213; Open in 6,020,539 grades.Can used according to the inventionly be suitable for transgenosis is inserted any methods for plant transformation of specified plant.
According to the present invention, the polynucleotide of importing can stably be kept in vegetable cell, if it is incorporated achromosomal autonomy replicon into or is integrated in the plant chromosome.Alternatively, the polynucleotide of importing may reside on the extrachromosomal nonreplication vector and can express instantaneously or have instantaneous activity.
The present invention also is embodied in to produce and comprises in the method for the transgenic plant of listed at least one polynucleotide in the table 1, when wherein comparing with the wild-type kind of this plant, the expression of described polynucleotide in this plant causes growth and/or the output of this plant under normal and/or the condition of restricting water supply to increase and/or the environmental stress-tolerance raising, described method comprises step: (a) expression cassette mentioned above is imported vegetable cell, (b) from plant transformed cell regeneration transgenic plant; And select the higher plant of output from the regenerated vegetable cell.This vegetable cell includes but not limited to protoplastis, the cell that produces gamete and the cell of the complete plant of regeneration.As used herein, term " genetically modified " refers to contain any plant, vegetable cell, callus, plant tissue or the plant part of expression cassette mentioned above.According to the present invention, this expression cassette is integrated into karyomit(e) or to the stable extra-chromosomal element with being stabilized, thereby it is passed to the follow-up generation.
Genetic modification can cultivate the plant of improvement to the influence of plant-growth and/or output and/or stress tolerance under normal and/or inferior suitable condition and growth characteristics and/or the metabolism of this plant of subsequent analysis assessed.This type of analytical technology is well known to those skilled in the art, and comprise measure dry weight, weight in wet base, seed weight, number seeds, polypeptide is synthetic, sugar is synthetic, lipid is synthetic, evapotranspiration speed, overall plant and/or crop yield, bloom, breed, tie kind, root growth, respiratory rate, photosynthesis rate, metabolite composition etc.
The present invention is further specified by following examples, and described embodiment should not be interpreted as scope of the present invention is limited by any way.
Embodiment 1
The sign of gene
Use standard recombinant technology has been cloned gene B0821 (SEQ ID NO:1), B1187 (SEQ ID NO:15), B2173 (SEQ ID NO:17), B2668 (SEQ ID NO:3), B2670 (SEQ ID NO:21), B3362 (SEQ ID NO:5), B3555 (SEQ ID NO:7), SLL1911 (SEQ ID NO:9), SLR1062 (SEQ ID NO:11), YBR222C (SEQ ID NO:23), YDL193W (SEQ ID NO:13), YNL030W (SEQ ID NO:39), YLR251W (SEQ ID NO:45), YPR036W (SEQ ID NO:57), SLL1326 (SEQ ID NO:61), SLR1329 (SEQ ID NO:65), SLR0977 (SEQ ID NO:67), ssr0390 (SEQ ID NO:69), sll1382 (SEQ ID NO:71), sll0248 (SEQ ID NO:77), sll0819 (SEQ ID NO:79), sll1796 (SEQ ID NO:89), slr1739 (SEQ ID NO:91), sll0378 (SEQ ID NO:93), slr1368 (SEQ ID NO:95) and sll0099 (SEQ ID NO:97).The aminoacid sequence of the prediction by more described gene and the function that known other genes of function are predicted each gene.Use currently known methods, separate homologue cDNA from the patent library of each species.Use the bioinformatic analysis method to handle and the note sequence.In selecting, uses homologous sequence as described below institute's separation sequence and the amino acid identity and the similarity degree of close known common sequence accordingly.Use Paired comparison method: gap penalty: 11; Point penalty is extended in the room: 1; Matrix: blosum62 keeps the score.
B2173 (SEQ ID NO:17) is a Nucleotide binding domains protein gene.With the predicted amino acid sequence of the total length of this gene at the patent database of the canclin sequence of prediction with e value e -10Carry out blast search (above Altschul etc.).Identify respectively a homologue from soybean and corn.The amino acid dependency of these sequences shows in the comparison result shown in Fig. 1.
The full length DNA sequence of YBR222C (SEQ ID NO:23) coding is from the peroxysome-coenzyme A synthetic enzyme of yeast saccharomyces cerevisiae.With the predicted amino acid sequence of the total length of this gene at the patent database of canola oil dish, soybean, rice and corn cDNA with e value e -10Carry out blast search (above Altschul etc.).Identify from 3 homologues of canola oil dish with from 4 homologues of soybean.The amino acid dependency of these sequences shows in the comparison result shown in Fig. 2.
The full length DNA sequence of YNL030W (SEQ ID NO:39) coding is from the histone H 4 of yeast saccharomyces cerevisiae.With the predicted amino acid sequence of the total length of this gene at the patent database of rice and flax cDNA with e value e -10Carry out blast search (above Altschul etc.).Identify respectively a homologue from rice and flax.The amino acid dependency of these sequences shows in the comparison result shown in Fig. 3.
YLR251W (SEQ ID NO:45) is a SYM1 type conformity membrane albumen.With the predicted amino acid sequence of the total length of this gene at the patent database of the predicted amino acid sequence of canola oil dish, barley, soybean, flax and rice with e value e -10Carry out blast search (above Altschul etc.).Identify a homologue from each library.The amino acid dependency of these sequences shows in the comparison result shown in Fig. 4.
YPR036W (SEQ ID NO:57) is a vacuolar proton pump subunit H albumen.With the predicted amino acid sequence of the total length of this gene at the patent database of the predicted amino acid sequence of canola oil dish with e value e -10Carry out blast search (above Altschul etc.).Identify a homologue from the canola oil dish.The amino acid dependency of these sequences shows in the comparison result shown in Fig. 5.
SLL1326 (SEQ ID NO:61) is an atp synthase subunit α albumen.With the predicted amino acid sequence of the total length of this gene at the patent database of the aminoacid sequence of prediction with e value e -10Carry out blast search (above Altschul etc.).Identify a homologue from flax library.The amino acid dependency of these sequences shows in the comparison result shown in Fig. 6.
Ferredoxin in sll1382 (SEQ ID NO:71) the genes encoding cytoalgae.With the full length amino acid sequence of sll1382 at the cDNA patent database with e value e -10Carry out blast search (above Altschul etc.).Identify from 1 homologue of canola oil dish with from 1 homologue of soybean.The amino acid dependency of these sequences shows in the comparison result shown in Fig. 7.
Photosystem I reactive center subunit III in sll0819 (SEQ ID NO:79) the genes encoding cytoalgae.With the full length amino acid sequence of sll0819 at the cDNA patent database with e value e -10Carry out blast search (above Altschul etc.).Identify from 2 homologues of canola oil dish with from 2 homologues of soybean.The amino acid dependency of these sequences shows in the comparison result shown in Fig. 8.
Embodiment 2
The overexpression of selected gene in plant
Use currently known methods, the polynucleotide of table 1 are connected in the expression cassette.Use 3 different promotors to control the expression of transgenosis in Arabidopis thaliana: the gene or SEQ ID NO:105, super promotor (SEQ ID NO:103) and the parsley ubiquitin promotor (SEQ ID NO:102) that are used to express from intestinal bacteria and cyanobacteria from the USP promotor (SEQ ID NO:104) of broad bean are used to express the gene from yeast saccharomyces cerevisiae.For the selectivity target of polypeptide, use in table 8,9,12,13,15-18,20-25 and 27, be called " Mito " from the mitochondrial transport peptide in the arabidopsis gene of coding line plastochondria isovaleryl-CoA dehydrogenase.SEQ ID NO:107 is used to express from the gene of intestinal bacteria and cyanobacteria or SEQ ID NO:109 and is used to express gene from yeast saccharomyces cerevisiae.In addition, for orientation expression, use the chloroplast transit peptides (SEQ ID NO:111) of spinach (Spinacia oleracea) gene that in table 6,14,16,17,19-23 and 25, is called the coding ferredoxin-nitrite reductase of " Chlor ".
Use currently known methods, with the environmental C24 of the construct arabidopsis thaliana transformation that contains gene described in the embodiment 1.Based on the type (chloroplast(id), mitochondrial and non-target-latter means and do not add extra target signal) that drives expression promoter, gene source species and target, compile the seed that transforms plant from T2.Seed compiles under the growth conditions that thing is used for fully watering and restricting water supply to be chosen at the primary screen of biomass.Selecting primary screen chooses from the hit results of compiling thing, carries out analysis of molecules and collect seed.The seed of collecting is used for the analysis of secondary screens subsequently, in secondary screens at each transgenic event more the individuality of high number analyze.If identify in the secondary screens that the plant from certain construct has the increase biomass compared with the control, then this plant experiences three screening.In this screening, fully water and arid growth conditions under measure the 100 strain plants that surpass from whole transgenic events of this construct.Use canonical statistics method, from the data of described transgenic plant and wild-type arabidopsis thaliana or with the plant comparison of from the transgenic arabidopsis seed of a collection of selection at random, cultivating.
Water to soil saturation for twice weekly to fully watering the plant of cultivating under the water condition.Use commercial imaging system, took the image of transgenic plant on the 17th and the 21st.Alternatively, by frequently not watering and cultivate plants under the growth conditions of restricting water supply to soil saturation (this causes that soil becomes dry watering between the water treatment).In these experiments, gave water at after planting the 0th, 8 and 19 day.Use commercial imaging system, took the image of transgenic plant on the 20th and the 27th.
Use image analysis software to come the transgenosis of cultivating in the comparison identical experiment and the image of control plant.Described image is used for the relative size of plant or biomass definite pixel and color as plant are defined as the ratio of deep green to the total area.The ratio in back (health index by name) be in the leaf chlorophyll relative quantity measure and thereby be measuring of leaf aging or flavescence relative quantity, and only at the 27th day entry.Containing existence variation between the transgenic plant of several genes, reason is that DNA inserts site difference and other factors that influence gene expression dose or pattern.
Table 2 shows the comparative result of the value of arabidopsis thaliana to table 27.Percent change is represented the value of transgenic plant with respect to control plant, the percentage ratio of non-transgenic plant in contrast; The p value is to check the difference statistical significance of comparative result between transgenosis and the control plant based on the T of whole independent eventss, and wherein NS is illustrated on 5% probability level not remarkable; Event number is represented the sum of the independent transgenic event checked in this experiment; Positive events is represented the sum greater than the independent transgenic event of contrast in this experiment; Negative representations of events is less than the sum of the independent transgenic event of contrast in this experiment.NS is illustrated on 5% probability level not remarkable.
A. the agnoprotein matter of non-target
The protein that is called B0821 (SEQ ID NO:2) is expressed in Arabidopis thaliana, wherein uses B0821 to express to be subjected to super promotor (super promoter) to control and does not have a construct that external source target sequence is added into SEQ ID NO:2.Table 2 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with these constructs and detect under the condition of restricting water supply.
Table 2
Figure BDA0000062684420000481
The arabidopsis thaliana that table 2 is presented at the expression B0821 (SEQ ID NO:2) that cultivates under the condition of restricting water supply is obviously bigger than the control plant of not expressing B0821 (SEQ ID NO:2) on 27th.Table 2 shows that also most independent transgenic event comparison is according to bigger.
Use and wherein transcribe the construct that is subjected to super promotor control, in Arabidopis thaliana, express B2668 gene (SEQ ID NO:4), the protein of this genes encoding Unknown Function.Table 3 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with these constructs and detect under the condition of restricting water supply.
Table 3
It is obviously bigger according to plant that table 3 is presented at 2 experimental comparison of arabidopsis thaliana in 3 experiments of cultivating under the condition of restricting water supply.Table 3 shows that also most independent transgenic event comparison is according to bigger.
Use and wherein transcribe the construct that is subjected to super promotor control, in Arabidopis thaliana, express B3362 gene (SEQ ID NO:6), the protein of this genes encoding Unknown Function.Table 4 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with this construct and detect under the condition of restricting water supply.
Table 4
Table 4 shows when cultivating that the arabidopsis thaliana comparison of expressing B3362 (SEQ ID NO:6) is obviously bigger according to plant under plant is being restricted water supply condition.Table 4 shows that also most independent transgenic event comparison is according to bigger.In addition, this construct is increased in the amount of the green color of plant when cultivating under the condition of restricting water supply significantly.
Use and wherein transcribe the construct that is subjected to super promotor control, in Arabidopis thaliana, express B3555 gene (SEQ ID NO:8), the protein of this genes encoding Unknown Function.Table 5 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with this construct and detect under the condition of restricting water supply.
Table 5
Figure BDA0000062684420000511
Table 5 shows when cultivating under plant is being restricted water supply condition, expresses the arabidopsis thaliana of B3555 (SEQ ID NO:8) and compares according to plant obviously bigger generally.Table 5 shows that also most independent transgenic event comparison is according to bigger.In addition, when comparing with the SuperPool contrast, this construct is increased in the amount of the green color of plant when cultivating under the condition of restricting water supply significantly.
B. the agnoprotein matter of plastid target
Use and wherein transcribe two constructs that are subjected to the control of PcUbi promotor, in Arabidopis thaliana, express SLL1911 gene (SEQ ID NO:10), the protein of this genes encoding Unknown Function.In a construct, chloroplast targeted peptide is connected with SEQ ID NO:10 effectively, and another construct does not have exogenous target peptide.Table 6 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with this construct and detect under the condition of restricting water supply.
Table 6
Figure BDA0000062684420000521
Table 6 shows that the arabidopsis thaliana comparison photograph plant of expressing SLL1911 (SEQ ID NO:10) is obviously bigger when cultivating under SLL1911 target chloroplast(id) and plant are being restricted water supply condition.Table 6 shows also that when SLL1911 target chloroplast(id) most independent transgenic event comparison is according to bigger.The amount of the green color of plant when in addition, wherein the construct that effectively is connected with SLL1911 of the chloroplast targeted peptide of external source is increased in cultivation under the condition of restricting water supply significantly.These data show when SLL1911 effectively is connected with chloroplast targeted peptide, and described plant is compared the photograph plant and produces more chlorophyll or have still less chlorophyll degradation during coercing.On the contrary, when expression of plants lacked the SLL1911 form of the chloroplast targeted peptide of external source, with the control plant of cultivating under the identical condition of restricting water supply relatively the time, the transgenic plant of gained were littler and have a significantly less green color.Generally speaking, the Subcellular Localization of these observations promptings SLL1911 is important to the size of the transgenic plant of increase expression SLL1911 gene and the amount of green color.
Use and wherein transcribe the construct that is subjected to the control of PcUbi promotor, in Arabidopis thaliana, express SLR1062 gene (SEQ ID NO:12), the protein of this genes encoding Unknown Function.Table 7 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with this construct and detect under the condition of restricting water supply.
Table 7
Figure BDA0000062684420000531
Figure BDA0000062684420000541
Table 7 shows when cultivating under plant is being restricted water supply condition, expresses the arabidopsis thaliana of SLR1062 (SEQ ID NO:12) and compares according to plant obviously bigger usually.Table 7 shows that also most independent transgenic event comparison is according to bigger.In addition, in two observations in 3 observations, this construct is increased in the amount of the green color of plant when cultivating under the condition of restricting water supply significantly.These data show that described plant is compared according to plant and produce more chlorophyll or have still less chlorophyll degradation during coercing.
C. 11 isoprene pyrophosphate synthetases
Use and wherein transcribe the effective construct that is connected with Mitochondrially targeted peptide of the polypeptide that is subjected to the control of USP promotor and from the gained transcript, translates, the YDL193W gene (SEQ ID NO:14) of the 11 isoprene pyrophosphate synthetases that expressing encodes infers in Arabidopis thaliana.Table 8 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with this construct and detect under the condition of restricting water supply.
Table 8
Figure BDA0000062684420000542
Table 8 shows when cultivating that the arabidopsis thaliana comparison of expressing YDL193W (SEQ ID NO:14) is obviously bigger according to plant under plant is being restricted water supply condition.Table 8 shows that also most independent transgenic event comparison is according to bigger.In addition, this construct is increased in the amount of the green color of plant when cultivating under the condition of restricting water supply significantly.The greener color of volume shows that described plant is compared according to plant and produces more chlorophyll or have still less chlorophyll degradation during coercing.
D. the transcriptional of inferring of fatty acid metabolism
Use and wherein transcribe the mitochondrial construct of transcriptional target that is subjected to control of USP promotor and fatty acid metabolism, in Arabidopis thaliana, express the transcriptional of inferring of the fatty acid metabolism that is called B1187 (SEQ ID NO:16).Table 9 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with these constructs and detect under fully watering water condition.
Table 9
Figure BDA0000062684420000551
It is obviously bigger than the control plant of not expressing B1187 (SEQ ID NO:16) that table 9 is presented at the arabidopsis thaliana that fully waters cultivation under the water condition.Table 9 also shows whole independent transgenic events, and all the comparison photograph is bigger in the environment that fully waters.
E. the nucleotide binding protein that has the P of G3E family ring structure territory
Use and wherein transcribe the construct that is subjected to super promotor control, express B2173 gene (SEQ ID NO:18) in Arabidopis thaliana, this genes encoding has the nucleotide binding protein in the P of G3E family ring structure territory.Table 10 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with these constructs and detect under the condition of restricting water supply.
Table 10
Table 10 shows that the arabidopsis thaliana of expressing B2173 (SEQ ID NO:18) is obviously bigger than SuperPool control plant.Table 10 shows that also most independent transgenic event is bigger than the SuperPool contrast.
The membranin of F. inferring
Use and wherein transcribe the construct that is subjected to super promotor control, in Arabidopis thaliana, express B2670 gene (SEQ ID NO:22), the membranin that this genes encoding is inferred.Table 11 has been described biomass and the health index data that obtain from the arabidopsis thaliana of using two construct conversions and detecting under the condition of restricting water supply.
Table 11
Figure BDA0000062684420000571
Table 11 is presented at when cultivating under the condition of restricting water supply, and the arabidopsis thaliana comparison of expressing B2670 (SEQ ID NO:22) is obviously bigger according to plant.In addition, these transgenic plant are compared on color according to showing darker green.These data show that described plant is compared according to plant and produce more chlorophyll or have still less chlorophyll degradation during coercing.Table 11 shows that also most independent transgenic event comparison is according to bigger.
G. peroxysome-coenzyme A synthetic enzyme
Use and wherein transcribe the effective construct that is connected with Mitochondrially targeted peptide of the polypeptide that is subjected to the control of USP promotor and from the gained transcript, translates, in Arabidopis thaliana, express the YBR222C gene (SEQ ID NO:24) of encoding superoxide enzyme body coenzyme A synthetic enzyme.Table 12 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with this construct and detect under the condition of restricting water supply.
Table 12
Figure BDA0000062684420000581
Table 12 shows when cultivating that the arabidopsis thaliana comparison of expressing YBR222C (SEQ ID NO:24) is obviously bigger according to plant under plant is being restricted water supply condition.Table 12 shows that also most independent transgenic event comparison is according to bigger.In addition, this construct is increased in the amount of the green color of plant when cultivating under the condition of restricting water supply significantly.The greener color of volume shows that described plant is compared according to plant and produces more chlorophyll or have still less chlorophyll degradation during coercing.
H. histone H 4
Use and wherein transcribe the effective construct that is connected with Mitochondrially targeted peptide of the polypeptide that is subjected to the control of USP promotor and from the gained transcript, translates, in Arabidopis thaliana, express the YNL030W gene (SEQ ID NO:40) of coding histone H 4.Table 13 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with this construct and detect under fully watering water condition.
Table 13
Figure BDA0000062684420000591
Table 13 shows when plant is fully watered, and expresses the arabidopsis thaliana of YNL030W (SEQ ID NO:40) and compares according to plant obviously bigger generally.Table 13 shows that also most independent transgenic event comparison is according to bigger.In addition, under fully watering water condition, cultivate and when comparing with the MTXC24 contrast, this construct increases the amount of the green color of plant significantly.The greener color of volume shows that described plant is compared according to plant and produces more chlorophyll or have still less chlorophyll degradation during coercing.
I. conformity membrane Protein S YM1
Use SYM1 type conformity membrane protein expression wherein to be subjected to the construct of USP promotor, super promotor or control of PCUbi promotor and conformity membrane targeting proteins chloroplast(id), in Arabidopis thaliana, express the conformity membrane albumen that is called YLR251W (SEQ ID NO:45).Table 14 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with these constructs and detect under (WW) condition of restricting water supply (CD) and fully water.
Table 14
Figure BDA0000062684420000601
Figure BDA0000062684420000611
Table 14 be presented at promotor PCUbi (SEQ ID NO:102) or USP (SEQ ID NO:104) control express down transgenic plant that YLR251W (SEQ ID NO:62) gene is targeted to plastid fully water or drought condition under obviously bigger than the control plant of not expressing YLR251W (SEQ ID NO:45) gene.In these experiments, whole or most independent transgenic event with these two promotors is compared under the round-robin drought environment according to bigger.Compare under the round-robin drought environment according to bigger observations as transgenic plant and to confirm, when use USP or PCUbi promoter expression, the existence of SYM1 albumen in plastid causes the harmful effect that water is lost that is attributed to of the transport efficacy improved and attenuating.
Table 14 be presented at the control of super promotor express down transgenic plant that YLR251W (SEQ ID NO:45) gene is targeted to plastid fully water or drought condition under obviously littler than the control plant of not expressing YLR251W (SEQ ID NO:45) gene.The function that the YLR251W (SEQ ID NO:45) that these results show to be provided by PCUbi and USP expresses for YLR251W (SEQ ID NO:45) is important.
J. vacuolar proton pump subunit H
Use wherein vacuolar proton pump subunit H protein expression to be subjected to control of USP promotor and the mitochondrial construct of vacuolar proton pump subunit H targeting proteins, in Arabidopis thaliana, express the vacuolar proton pump subunit H albumen that is called YPR036W (SEQ ID NO:58).Table 15 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with these constructs and detect under fully watering water condition.
Table 15
Figure BDA0000062684420000621
Table 15 is presented at USP promotor control and expresses YPR036W (SEQ ID NO:58) gene down to be targeted to mitochondrial transgenic plant obviously bigger and more healthy than the control plant of not expressing YPR036W (SEQ ID NO:58) gene under drought condition.These the experiment in, most have Mitochondrially targeted independent transgenic event under the round-robin drought environment, compare the photograph bigger and more healthy.Compare under the round-robin drought environment according to bigger and more healthy observations as transgenic plant and to confirm, the existence of V-type ATP enzyme subunit H albumen in plastosome causes the harmful effect that water is lost that is attributed to of the transport efficacy improved and attenuating.
K.F-ATP enzyme subunit α
F-ATP enzyme subunit α gene SLL1326 (SEQ ID NO:62) expresses and target plastid and plastosome or plastid in Arabidopis thaliana under the control of PCUbi promotor.Table 16 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with these constructs and detect under the round-robin drought condition.
Table 16
Figure BDA0000062684420000631
Table 16 is presented at PCUbi promotor control and expresses the SLL1326 gene down to be targeted to mitochondrial transgenic plant obviously bigger than the control plant of not expressing the SLL1326 gene under drought condition.These the experiment in, most have Mitochondrially targeted independent transgenic event under the round-robin drought environment, compare the photograph bigger.Compare under the round-robin drought environment according to bigger observations as transgenic plant and to confirm, the existence of F-ATP enzyme subunit α albumen in plastosome causes the harmful effect that water is lost that is attributed to of the transport efficacy improved and attenuating.
Table 16 is presented at PCUbi promotor control and expresses the SLL1326 gene down to be targeted to the transgenic plant of plastid obviously littler and more unhealthy than the control plant of not expressing the SLL1326 gene under drought condition.Table 16 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with these constructs and detect under the condition of restricting water supply.
L.F-ATP enzyme subunit β
F-ATP enzyme subunit β gene SLR1329 (SEQ ID NO:66) expresses and target plastid or plastosome in Arabidopis thaliana under the control of PCUbi promotor.Table 17 has been described from transforming with these constructs and in the round-robin arid or fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 17
Figure BDA0000062684420000641
Table 17 is presented at PCUbi promotor control and expresses SLR1329 (SEQ ID NO:66) gene down to be targeted to mitochondrial transgenic plant obviously bigger and more healthy than the control plant of not expressing SLR1329 (SEQ ID NO:66) gene under drought condition.In these experiments, most independent transgenic event with plastosome guiding is compared under the round-robin drought environment according to bigger.Compare under the round-robin drought environment according to bigger observations as transgenic plant and to confirm, the existence of F-ATP enzyme subunit β albumen in plastosome causes the harmful effect that water is lost that is attributed to of the transport efficacy improved and attenuating.
Table 17 is presented at PCUbi promotor control and expresses transgenic plant that SLR1329 (SEQ ID NO:66) gene is targeted to plastid down in arid with fully water under the water condition obviously forr a short time than the control plant of not expressing SLR1329 (SEQ ID NO:66) gene, contrasts more unhealthy significantly under drought condition.
The M.ABC translocator
ABC transporter gene SLR0977 (SEQ ID NO:68) expresses and the target plastosome in Arabidopis thaliana under the control of PCUbi promotor.Table 18 has been described from transforming with this construct and in the round-robin arid with fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 18
Figure BDA0000062684420000651
Table 18 is presented at PCUbi promotor control and expresses the SLR0977 gene down to be targeted to mitochondrial transgenic plant obviously bigger than the control plant of not expressing the SLR0977 gene under drought condition.In these experiments, whole or most independent transgenic event with plastosome guiding is compared under the round-robin drought environment according to bigger.Compare under the round-robin drought environment according to bigger observations as transgenic plant and to confirm, the existence of abc transport albumen in plastosome causes the harmful effect that water is lost that is attributed to of the transport efficacy improved and attenuating.
N.PsaK
PsaK gene SSR0390 (SEQ ID NO:69) expresses and the target plastid in Arabidopis thaliana under the control of PcUbi promotor.Table 19 has been described biomass and the health index data that obtain from the arabidopsis thaliana that transforms with these constructs and detect under the round-robin drought condition.
Table 19
Figure BDA0000062684420000652
Figure BDA0000062684420000661
Table 19 show to express transgenic plant that the ssr0390 gene is targeted to plastid fully water under the water condition obviously bigger than the control plant of not expressing the ssr0390 gene.In these experiments, most independent transgenic event with plastid guiding is compared under the round-robin drought environment according to bigger.Compare under the round-robin drought environment according to bigger observations as transgenic plant and to confirm, the existence of PsaK albumen in plastid causes the harmful effect that water is lost that is attributed to of the photosynthetic efficiency improved and attenuating.
O. ferredoxin (PetF)
(construct is under the control of PcUbi promotor and the target plastosome to use two different constructs, and second construct has identical promotor and target plastid), in Arabidopis thaliana, express ferredoxin (PetF) gene sll1382 (SEQ ID NO:71).Table 20 has been described from transforming with these constructs and in the round-robin arid with fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 20
Figure BDA0000062684420000662
Table 20 show to express the sll1382 gene be targeted to mitochondrial transgenic plant fully water under the water condition obviously bigger than the control plant of not expressing the sll1382 gene.Under the condition of restricting water supply, when measuring in 27th, transgenic plant comparison illumination shows littler, and does not have obvious difference on other Measuring Time points or aspect health index.
The transgenic plant that table 20 demonstration expression sll1382 gene is targeted to plastid are obviously littler than the control plant of not expressing the sll1382 gene under the condition of restricting water supply.Extraly, with respect to the contrast under the condition of restricting water supply, these transgenic plant have lower health index scoring.Fully watering under the water condition, the transgenic plant that expression sll1382 gene is targeted to plastid do not have obviously different with contrast aspect biomass or health index.In these experiments, most independent transgenic event with plastosome guiding the two one of water surrounding under comparison according to bigger.
These observationss meet previous report, and described report shows that ferredoxin does not improve plant-growth when the plastid in the target transgenic plant.Confirm according to the bigger observations of plant in the time comparison of ferredoxin target plastosome that as transgenic plant the existence of ferredoxin in plastosome causes the electron transport efficient improved.
P. flavodoxin
Use two different constructs, flavodoxin gene sll0248 (SEQ ID NO:77) expresses and target plastosome or plastid in Arabidopis thaliana under the control of PcUbi promotor.Table 21 has been described from transforming with these constructs and in the round-robin arid with fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 21
Figure BDA0000062684420000671
Table 21 show to express transgenic plant that the sll0248 gene is targeted to plastid fully water under the water condition obviously bigger than the control plant of not expressing the sll0248 gene.Express sll0248 gene and ubcellular target to mitochondrial transgenic plant fully water under the water condition obviously littler on 17th than the control plant of not expressing the sll0248 gene, still 21 days with the same terms under do not express the sll0248 gene control plant do not have obviously different.Express the two one of the health index of transgenic plant of construct do not have obviously different with contrast.In these experiments, most independent transgenic event with plastid guiding the two one of water surrounding under comparison according to bigger, and those independent transgenic events with plastosome guiding under the environment that fully waters, compare shine littler.
Compare under the round-robin drought environment according to bigger observations as transgenic plant and to confirm, the existence of flavodoxin albumen in plastid causes the harmful effect that water is lost that is attributed to of the photosynthetic efficiency improved and attenuating.
Q.PsaF
Use two different constructs, PsaF gene SLL0819 (SEQ ID NO:79) expresses and target plastosome or target plastid in Arabidopis thaliana under the control of PcUbi promotor.Table 22 has been described from transforming with these constructs and in the round-robin arid with fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 22
Table 22 show to express transgenic plant that the ssr0390 gene is targeted to plastid fully water under the water condition obviously bigger than the control plant of not expressing the sll0819 gene.In these experiments, most independent transgenic event with plastid guiding is compared under the round-robin drought environment according to bigger.Compare under the round-robin drought environment according to bigger observations as transgenic plant and to confirm, the existence of PsaK albumen in plastid causes the harmful effect that water is lost that is attributed to of the photosynthetic efficiency improved and attenuating.
R.PetJ
Use two different constructs, PetJ gene SLL1796 (SEQ ID NO:89) expresses and target plastosome or target plastid in Arabidopis thaliana under the control of PcUbi promotor.Table 23 has been described from transforming with these constructs and in the round-robin arid with fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 23
Figure BDA0000062684420000691
It is obviously bigger than the control plant of not expressing the sll1796 gene under the condition of restricting water supply that table 23 shows that expression sll1796 gene is targeted to mitochondrial transgenic plant.Really have variation between the transgenic plant that contain the sll1796 gene, reason is DNA insertion site difference and other factors that influence gene expression dose or pattern.Health index is similar between transgenic plant and control plant.In these experiments, most independent transgenic event comparison is according to bigger.
The sll1796 gene is expressed in table 23 demonstration and ubcellular target to the transgenic plant of plastid are being restricted water supply and fully watering under the water condition obviously littler than the control plant of not expressing the sll1796 gene.In these experiments, whole independent transgenic event comparisons are according to littler.
Confirm according to the bigger observations of plant in the time comparison of PetJ targeting proteins plastosome that as transgenic plant the existence of PetJ albumen in plastosome causes the plastosome electron transport efficient improved.
S.PsbW
Use two different constructs, PsbW gene SLR1739 (SEQ ID NO:91) expresses and target plastosome or target plastid in Arabidopis thaliana under the control of PcUbi promotor.Table 24 has been described from transforming with these constructs and in the round-robin arid with fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 24
Figure BDA0000062684420000701
Table 24 demonstration is expressed the transgenic plant of slr1739 gene and is being restricted water supply and fully watering under the water condition obviously bigger than the control plant of not expressing the slr1739 gene.Health index round-robin arid and fully water transgenic plant under the water condition and control plant between be similar.In these experiments, most independent transgenic event the two one of water surrounding under comparison according to bigger.
Confirm according to the bigger observations of plant in the time comparison of PsbW targeting proteins plastosome as transgenic plant, the existence of PsbW albumen in plastosome cause fully water with drought condition under the electron transport efficient improved.
T.CobA(CysG)
Use two different constructs, uroporphyrin-III C-methyl transferase gene SLL0378 (SEQ ID NO:93) expresses and target plastosome or target plastid in Arabidopis thaliana under the control of PcUbi promotor.Table 25 has been described from transforming with these constructs and in the round-robin arid with fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 25
Figure BDA0000062684420000711
Table 25 demonstration expression sll0378 gene is targeted to the transgenic plant of plastid and is restricting water supply and fully watering under the water condition obviously bigger than the control plant of not expressing the sll0378 gene.In addition, the transgenic plant of cultivating under the condition of restricting water supply are compared on color according to having more deep green, shown in the health index that increases.This shows that described plant is compared according to plant and produces more chlorophyll or have still less chlorophyll degradation during coercing.
Table 25 shows that expression sll0378 gene is targeted to mitochondrial transgenic plant and is restricting water supply and fully watering under the water condition obviously littler than the control plant of not expressing the sll0378 gene.Extraly, with respect to the contrast under the condition of restricting water supply, these transgenic plant have the scoring of lower health index, but have higher health index scoring under the water condition fully watering.In these experiments, most independent transgenic event with plastid guiding the two one of environment under comparison according to bigger.
Bigger according to plant as transgenic plant in the time comparison of CobA targeting proteins plastid, but the observations bigger unlike control plant confirms when CobA targeting proteins plastosome, and the existence of CobA albumen in plastid causes the light collecting light ability that improves and shift to the more high efficiency energy of photosystem.
U. preceding corrin-8w decarboxylase (CbiT, CobL)
In Arabidopis thaliana, under PcUbi promotor control, express preceding corrin-8w decarboxylase gene Sll1368 (SEQ ID NO:95) during no ubcellular target.Table 26 has been described from transforming with these constructs and in the round-robin arid with fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 26
Figure BDA0000062684420000721
Table 26 shows that the transgenic plant of expressing the slr1368 gene are obviously bigger than the control plant of not expressing the slr1368 gene under the condition of restricting water supply.In addition, the transgenic plant of cultivating under the condition of restricting water supply are compared on color according to having more deep green, shown in the health index that increases.This shows that described plant is compared according to plant and produces more chlorophyll or have still less chlorophyll degradation during coercing.In these experiments, most independent transgenic event is compared under the environment of restricting water supply according to bigger.
Do not having obviously different with contrast aspect biomass or the health index fully watering the transgenic plant that the water condition following table reaches the slr1368 gene.
Confirm according to the bigger observations of plant that as transgenic plant comparisons the proteic existence of CbiT causes the light collecting light ability that improves and to the more high efficiency energy transfer of photosystem.
Corrin before the V decarboxylation-6y c5, and the 15-methyltransgerase (CobL, CbiE/CbiT)
Use two different constructs, corrin before the decarboxylase-6y c5,15-methyl transferase gene Sll0099 (SEQ ID NO:97) are expressed in Arabidopis thaliana under the control of PcUbi promotor and target plastosome or do not carry out target.Table 27 has been described from transforming with these constructs and in the round-robin arid with fully water biomass and the health index data that obtain the arabidopsis thaliana that detects under the water condition.
Table 27
Figure BDA0000062684420000731
Table 27 shows that expression sll0099 gene is targeted to mitochondrial transgenic plant and is restricting water supply and fully watering under the water condition obviously bigger than the control plant of not expressing the sll0099 gene.In addition, the transgenic plant of cultivating under the condition of restricting water supply are compared on color according to having more deep green, shown in the health index that increases.This shows that described plant is compared according to plant and produces more chlorophyll or have still less chlorophyll degradation during coercing.Express the sll0099 gene simultaneously and not the transgenic plant of target under the condition of restricting water supply, compare illumination and shows bigger and have higher health index and mark.In these experiments, most independent transgenic event the two one of environment under comparison according to bigger.
Confirm according to the bigger observations of plant that as transgenic plant comparisons the proteic existence of CobL causes the light collecting light ability that improves and to the more high efficiency energy transfer of photosystem.
Figure IDA0000062684490000011
Figure IDA0000062684490000021
Figure IDA0000062684490000031
Figure IDA0000062684490000041
Figure IDA0000062684490000051
Figure IDA0000062684490000061
Figure IDA0000062684490000071
Figure IDA0000062684490000091
Figure IDA0000062684490000101
Figure IDA0000062684490000121
Figure IDA0000062684490000131
Figure IDA0000062684490000141
Figure IDA0000062684490000151
Figure IDA0000062684490000171
Figure IDA0000062684490000181
Figure IDA0000062684490000191
Figure IDA0000062684490000201
Figure IDA0000062684490000211
Figure IDA0000062684490000221
Figure IDA0000062684490000231
Figure IDA0000062684490000251
Figure IDA0000062684490000261
Figure IDA0000062684490000271
Figure IDA0000062684490000281
Figure IDA0000062684490000291
Figure IDA0000062684490000301
Figure IDA0000062684490000311
Figure IDA0000062684490000321
Figure IDA0000062684490000331
Figure IDA0000062684490000341
Figure IDA0000062684490000351
Figure IDA0000062684490000361
Figure IDA0000062684490000381
Figure IDA0000062684490000391
Figure IDA0000062684490000401
Figure IDA0000062684490000411
Figure IDA0000062684490000421
Figure IDA0000062684490000431
Figure IDA0000062684490000451
Figure IDA0000062684490000471
Figure IDA0000062684490000481
Figure IDA0000062684490000491
Figure IDA0000062684490000501
Figure IDA0000062684490000511
Figure IDA0000062684490000521
Figure IDA0000062684490000531
Figure IDA0000062684490000551
Figure IDA0000062684490000561
Figure IDA0000062684490000571
Figure IDA0000062684490000581
Figure IDA0000062684490000591
Figure IDA0000062684490000621
Figure IDA0000062684490000631
Figure IDA0000062684490000641
Figure IDA0000062684490000651
Figure IDA0000062684490000661
Figure IDA0000062684490000691
Figure IDA0000062684490000701
Figure IDA0000062684490000711
Figure IDA0000062684490000731
Figure IDA0000062684490000741
Figure IDA0000062684490000751
Figure IDA0000062684490000761
Figure IDA0000062684490000781
Figure IDA0000062684490000811
Figure IDA0000062684490000821
Figure IDA0000062684490000831
Figure IDA0000062684490000841
Figure IDA0000062684490000861
Figure IDA0000062684490000881
Figure IDA0000062684490000891
Figure IDA0000062684490000901
Figure IDA0000062684490000911
Figure IDA0000062684490000921
Figure IDA0000062684490000931
Figure IDA0000062684490000951
Figure IDA0000062684490000971
Figure IDA0000062684490000981
Figure IDA0000062684490000991
Figure IDA0000062684490001011
Figure IDA0000062684490001021
Figure IDA0000062684490001031
Figure IDA0000062684490001051
Figure IDA0000062684490001061
Figure IDA0000062684490001081
Figure IDA0000062684490001091
Figure IDA0000062684490001101
Figure IDA0000062684490001111
Figure IDA0000062684490001121
Figure IDA0000062684490001131
Figure IDA0000062684490001141
Figure IDA0000062684490001151
Figure IDA0000062684490001161
Figure IDA0000062684490001171
Figure IDA0000062684490001181
Figure IDA0000062684490001191
Figure IDA0000062684490001201
Figure IDA0000062684490001221
Figure IDA0000062684490001231
Figure IDA0000062684490001241
Figure IDA0000062684490001251
Figure IDA0000062684490001271
Figure IDA0000062684490001281
Figure IDA0000062684490001291
Figure IDA0000062684490001301
Figure IDA0000062684490001351
Figure IDA0000062684490001361
Figure IDA0000062684490001371
Figure IDA0000062684490001381
Figure IDA0000062684490001391
Figure IDA0000062684490001401
Figure IDA0000062684490001411
Figure IDA0000062684490001421
Figure IDA0000062684490001431
Figure IDA0000062684490001441
Figure IDA0000062684490001451
Figure IDA0000062684490001461
Figure IDA0000062684490001471
Figure IDA0000062684490001481
Figure IDA0000062684490001491
Figure IDA0000062684490001501
Figure IDA0000062684490001511
Figure IDA0000062684490001521
Figure IDA0000062684490001531
Figure IDA0000062684490001541
Figure IDA0000062684490001551
Figure IDA0000062684490001561
Figure IDA0000062684490001571
Figure IDA0000062684490001581
Figure IDA0000062684490001591
Figure IDA0000062684490001601
Figure IDA0000062684490001611
Figure IDA0000062684490001631
Figure IDA0000062684490001641
Figure IDA0000062684490001651
Figure IDA0000062684490001661
Figure IDA0000062684490001671
Figure IDA0000062684490001681
Figure IDA0000062684490001691
Figure IDA0000062684490001701
Figure IDA0000062684490001711
Figure IDA0000062684490001721
Figure IDA0000062684490001731
Figure IDA0000062684490001741
Figure IDA0000062684490001751
Figure IDA0000062684490001761
Figure IDA0000062684490001771
Figure IDA0000062684490001781
Figure IDA0000062684490001791
Figure IDA0000062684490001801
Figure IDA0000062684490001811
Figure IDA0000062684490001821

Claims (9)

1. use the expression cassette transgenic plant transformed, described expression cassette comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding plastid transit peptides; Have the isolating polynucleotide of the photosystem I reactive center subunit III psaF polypeptide of PSI_PsaF characteristic sequence with coding, described PSI_PsaF characteristic sequence comprises and is selected from the 3rd to the 158th amino acids by SEQ ID NO:80; SEQ ID NO:82 the 43rd to the 217th amino acids; SEQ ID NO:84 the 46th to the 220th amino acids; SEQ ID NO:86 the 50th to the 224th amino acids; PSI_PsaF characteristic sequence in the group of forming with SEQ ID NO:88 the 50th to the 224th amino acids; Wherein these transgenic plant are compared with the wild-type plant of the same breed that does not comprise this expression cassette and show the output that increases.
2. described transgenic plant of claim 1, wherein said polypeptide comprises SEQ ID NO:82 the 1st to the 217th amino acids; SEQ ID NO:84 the 1st to the 220th amino acids; SEQ ID NO:86 the 1st to the 224th amino acids; SEQ ID NO:88 the 1st to the 224th amino acids.
3. the described transgenic plant of claim 1 further describe to being selected from the species in the group of being made up of corn, soybean, cotton, canola oil dish, rice, wheat or sugarcane.
4. seed, it isozygotys to the transgenosis that comprises expression cassette, and described expression cassette comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding plastid transit peptides; Have the isolating polynucleotide of the photosystem I reactive center subunit III psaF polypeptide of PSI_PsaF characteristic sequence with coding, described PSI_PsaF characteristic sequence comprises and is selected from the 3rd to the 158th amino acids by SEQ ID NO:80; SEQ ID NO:82 the 43rd to the 217th amino acids; SEQ ID NO:84 the 46th to the 220th amino acids; SEQ ID NO:86 the 50th to the 224th amino acids; PSI_PsaF characteristic sequence in the group of forming with SEQ ID NO:88 the 50th to the 224th amino acids.
5. the described seed of claim 4, wherein said polypeptide comprises SEQ ID NO:82 the 1st to the 217th amino acids; SEQ ID NO:84 the 1st to the 220th amino acids; SEQ ID NO:86 the 1st to the 224th amino acids or SEQ ID NO:88 the 1st to the 224th amino acids.
6. the described transgenic plant of claim 1 further describe to being selected from the species in the group of being made up of corn, soybean, cotton, canola oil dish, rice or wheat.
7. increase the method for plant biomass, the method comprising the steps of
A) transform the wild-type plant cell with comprising genetically modified expression cassette, described expression cassette comprises the isolating polynucleotide of the coding promotor that is in effective connection; The isolating polynucleotide of coding plastid transit peptides; Have the isolating polynucleotide of the photosystem I reactive center subunit III psaF polypeptide of PSI_PsaF characteristic sequence with coding, described PSI_PsaF characteristic sequence comprises and is selected from the 3rd to the 158th amino acids by SEQ ID NO:80; SEQ ID NO:82 the 43rd to the 217th amino acids; SEQ ID NO:84 the 46th to the 220th amino acids; SEQ ID NO:86 the 50th to the 224th amino acids; PSI_PsaF characteristic sequence in the group of forming with SEQ ID NO:88 the 50th to the 224th amino acids.
B) from plant transformed cell regeneration transgenosis plantlet; With
C) selection shows the transgenic plant of the output that increases.
8. the described seed of claim 3, wherein said polypeptide comprises SEQ ID NO:82 the 1st to the 217th amino acids; SEQ ID NO:84 the 1st to the 220th amino acids; SEQ ID NO:86 the 1st to the 224th amino acids or SEQ ID NO:88 the 1st to the 224th amino acids.
9. the described method of claim 8, wherein said plant is corn, soybean, cotton, canola oil dish, rice, wheat or sugarcane.
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